Thermodynamic dissection of the substrate-ribozyme interaction in the hammerhead ribozyme

The free energy of substrate binding to the hammerhead ribozyme was compared for 10 different hammerheads that differed in the length and sequence of their substrate recognition helices. These hammerheads were selected because neither ribozyme nor substrate oligonucleotide formed detectable alternate secondary structures. The observed free energies of binding varied from -8 to -24 kcal/mol and agreed very well with binding energies calculated from the nearest-neighbor free energies if a constant energetic penalty of DeltaG degreescore = +3.3 +/- 1 kcal/mol is used for the catalytic core. A set of substrates that contained a competing hairpin secondary structure showed weaker binding to the ribozyme by an amount consistent with the predicted free energy for hairpin formation. These thermodynamic conclusions permit the prediction of substrate binding affinities for ribozyme-substrate pairs of any helix length and sequence, and thus, should be very valuable for the rational design of ribozymes directed toward gene inactivation.     

Antibody responses measured by various serologic tests in pigs orally inoculated with low numbers of Toxoplasma gondii oocysts

OBJECTIVE: To follow antibody responses measured by various serologic tests in pigs orally inoculated with low (< or = 10 oocysts) numbers of Toxoplasma gondii oocysts. ANIMALS: 24, 2- to 3-month-old pigs. PROCEDURE: Pigs (n = 42) were inoculated orally with 10 (14 pigs) or 1 (28 pigs) infective oocysts, and 6 pigs served as uninoculated controls. Blood (serum) samples were obtained at 1- to 3-week intervals until euthanasia. At necropsy, the brain, heart, and tongue of pigs were bioassayed in mice and cats for isolation of T gondii. Modified agglutination test (MAT), using whole, fixed tachyzoites and mercaptoethanol; latex agglutination test (LAT); indirect hemagglutination test (IHAT); Sabin-Feldman dye test (DT); and ELISA were used to evaluate serologic responses to T gondii. RESULTS: T gondii was isolated from tissues of 13 of 14 pigs each fed 10 oocysts, 17 of 28 pigs each fed 1 oocyst, and 0 of 6 control pigs. 29 of 30 T gondii-infected pigs developed antibodies when measured by MAT, DT, and ELISA; the 1 seronegative-infected pig had been fed 10 oocysts and was euthanatized 69 days after inoculation. LAT detected antibodies in 26 of 30 T gondii-infected pigs. IHAT detected antibodies in 11 T gondii-infected pigs. CONCLUSION: MAT, DT, and ELISA were more sensitive serologic assays than LAT and IHAT for detecting antibodies induced by low numbers of T gondii in pigs.     

Effects of comsumption of high and low nicotine tobacco byManduca sexta (Lepidoptera: Sphingidae) on survival of gregarious endoparasitoidCotesia congregata (Hymenoptera: Braconidae)

The significance of nicotine in the three trophic level interaction involving tobacco (Nicotiana tabacum), the tobacco hornworm (Manduca sexta), and the parasitoidCotesia congregata was investigated in field plots of two varieties of tobacco which had about a 10-fold difference in their nicotine content. WhileM. sexta mortality, rates of parasitism byC. congregata, and the total number ofC. congregata larvae produced per host were similar on each of the two varieties, the number of parasitoids reaching adult-hood on the low nicotine treatment was nearly twice that on the high nicotine treatment. This difference was due to the significantly greater proportion of parasitoid larvae which failed to emerge from the host or that died prior to pupation after emerging from hosts which fed on the high nicotine variety. A greater proportion of larvae from hosts which fed on the low nicotine tobacco died as pupae. No treatment differences occurred for either sex of the parasitoid in individual dry weight, longevity, or pupal development time, except that female pupal duration was prolonged in the high nicotine treatment. These results support the suggestion that plant allelochemicals, which may function to provide plant resistance against pest herbivores, can be detrimental to natural enemies of the pest.Scientific article No. A4148 of the Maryland Agricultural Experiment Station, Department of Entomology. Research supported by NSF grant BSR-84-00614.     

Heart muscle cells acutely infected with Trypanosoma cruzi: characterization of electrophysiology and neurotransmitter responses

Primary cell cultures of mouse ventricular myocardium were infected with Trypanosoma cruzi, to study the consequences of T. cruzi-muscle cell interaction on the rate of spontaneous contractions, on the responses to norepinephrine, and on action potential parameters. Single cells or small cell groups of infected cultures were subjected to pharmacological and electrophysiological experiments. In concentrations ranging from 1 nM to 100 microM, norepinephrine exerted positive or negative chronotropic effects mediated by alpha-adrenergic receptors. A significant number of infected cells (25%) did not respond to the agonist. Two days after infection the cultures exhibited a higher frequency of spontaneous contractions (20%), paralleled by an increase in firing rate and a decrease in the action potential duration without significant changes in maximum diastolic potential and action potential amplitude. A decrease in alpha-adrenergic receptor-mediated positive chronotropic response to norepinephrine was also observed in 2-day infected cells. Cells made to phagocyte ferritin particles showed an increase in the rate of spontaneous contractions, but no changes in the positive chronotropic responses to norepinephrine. In conclusion, these observations show that during acute infection with T. cruzi, there are alterations in automaticity and in the chronotropic responses to norepinephrine, whose mechanisms are related to the process of parasite endocytosis by the cardiac cells.     

A simplified approach to radionuclide venography: concise communication

A simplified method has been developed for performing radionuclide venography. The method makes use of the scintillation camera and a synchronized whole-body scanning bed. This technique permits a more integrated presentation of the data and is performed in conjunction with a standard ventilation-perfusion lung study. The total amount of 99mTc tracer injected is 2 mCi.     

Effects of a single contraceptive Silastic implant containing nomegestrol acetate on ovarian function and cervical mucus production during 2 years

OBJECTIVE: To study the mechanism of action of Uniplant (South to South Corporation in Reproductive Health, Salvador, Brazil), a single Silastic capsule containing nomegestrol acetate (Lutenyl, Theramex, France) in women during 2 years. DESIGN: Comparison between the hormonal levels, follicular development, cervical mucus (CM) production, and endometrial thickness in the menstrual cycle before implant insertion and at 1, 6, 12, 18, and 24 months after implant insertion. PARTICIPANTS: A total of 15 women of reproductive age were enrolled for the 1st year of use. Twelve of these women continued for a 2nd year of Uniplant use. MAIN OUTCOME MEASURES: Hormonal plasma levels were measured in control cycles and at 1, 6, 12, 18, and 24 months of Uniplant use. Cervical mucus, follicular development, and endometrial thickness also were evaluated. RESULTS: In this study, Uniplant blocks ovulation in 86 percent of cycles studied. Disturbances in follicular growth were observed also. Cervical mucus was scanty and viscous in all women during this study. Endometrial thickness was <8 mm in all cycles studied. CONCLUSION: This study shows that Uniplant is a long-acting contraceptive that probably acts at the hypothalamic-pituitary levels, on the ovary, on CM production, and on the endometrium. These properties suggest the use of Uniplant as a contraceptive agent, especially if one considers the lack of androgenic and metabolic effects and the maintenance of periodic bleeding similar to menstruation.