Determination of the biological activity of Clostridium difficile toxins in in vivo and in vitro experiments

The biological activity of the filtrates of 29 C. difficile strains was studied in vivo (suckling white mice) and in vitro (cell cultures of different species and origin). The action of the filtrates on the experimental models in vivo was evaluated from the cytotoxic effect index, while in vitro the intensity of the cytotoxic effect was evaluated from the percentage of dead cells in the monolayer. The results of the comparative determination of toxicity characteristics in vivo and in vitro demonstrated that cell cultures were more sensitive experimental models than suckling white mice. The use of cell cultures permitted the quantitative evaluation of the cytotoxic activity of the filtrates under study, as well as the detection of their cell-directed action at minimal concentrations.     

Partial deficiency of hypoxanthine-guanine phosphoribosyltransferase with reduced affinity for PP-ribose-P in four related males with gout

Summary A family is described in which four affected males, spanning two generations, have hyperuricemia and gout accompanied by hematuria but are without severe neurologic involvement. The affected males were found to have markedly reduced levels of erythrocytic hypoxanthine-guanine phosphoribosyltransferase (HGPRT) activity; these were 5–12% with hypoxanthine and 0.5–3% with guanine as compared to controls. Erythrocytic adenine phosphoribosyltransferase (APRT) was approximately three-fold elevated in the affected individuals.The residual HGPRT activity in affected males enabled characterization of some of the properties of this mutation. The apparent Michaelis constants (km) for both hypoxanthine and guanine were essentially unchanged, whereas the km for PP-ribose-P was approximately 10–20-fold elevated for all four affected males. The enzyme was more sensitive to product inhibition by IMP and GMP than controls, and exhibited greater thermal lability at 65°C than found with control lysates.     

Influence of carbachol and isoproterenol on myoelectrical activity of the uterus, vagina, and duodenum in the conscious domestic hen

The influence of intravenous infusion (20 min) of carbachol (CAR) and isoproterenol (IPN) on myoelectrical activity of the uterus, vagina, and duodenum was studied in 6 conscious and unrestrained hens. IPN infusions (1.5, 7.5, and 15 micrograms/hen/min) were performed before oviposition, and CAR infusions (0.5, 1.5, and 5 micrograms/hen/min) were administered after oviposition. CAR infusions resulted in a significant stimulation of myoelectrical activity of the vagina (1.5 and 5 micrograms/min) and the duodenum (5 micrograms/min), and in a significant increase (45 +/- 6%) in heart rate (HR), as compared to control values. These effects were blocked by previous i.v. injections of atropine (0.4 mg/hen). IPN infusions induced a significant inhibition of myoelectrical activity of the middle (7.5 and 15 micrograms/min) and caudal (all doses) uterine segments, of the vagina (7.5 and 15 micrograms/min), and of the duodenum (15 micrograms/min), and a significant tachycardia (39 +/- 14%) as compared to control values. The inhibitory effects of IPN on all organs studied and the stimulatory action on HR were suppressed by propranolol (0.24 mg/hen). The sensitivity of the vagina to both drugs argues for a more active participation of this oviductal part in the mechanism of oviposition.     

Prey-capture techniques and prey preferences of Corythalia canosa and Pystira orbiculata, ant-eating jumping spiders (Araneae, Salticidae)

Corythalia canosa from Florida is an unusual salticid because it is known to eat ants. This species' specialized behaviour for catching ants is described in detail for the first time and compared to its behaviour for catching other insects. Pystira orbiculata from Queensland is shown to be another ant-eating salticid, although its behaviour for catching ants seems less specialized than that of C. canosa. Three different types of tests of prey preference were carried out. In each type of test C. canosa and P. orbiculata took ants in preference to other insects. Another species of salticid, Trite planiceps from New Zealand, failed to eat ants in these tests, although T. planiceps often attacked then released the ants. Corythalia canosa's and P. orbiculata's preference for ants, and their prey-specific predatory behaviour for catching ants, are shown not to depend on prior experience with ants.     

Gene expression changes associated with cytotoxicity identified using cDNA arrays

In order to investigate gene expression changes associated with cytotoxicity, we used cDNA arrays to monitor the expression of over 5,000 genes in response to toxic stress in the HepG2 liver cell line. Cells were treated with cytotoxic doses of acetaminophen, caffeine or thioacetamide for nine time points ranging from 1 to 24 h. Samples of mRNA from each time point were used to prepare radiolabeled cDNA, which was hybridized to nylon-membrane-based cDNA arrays. High-stringency washes were applied to reduce cross-hybridization. Analysis of spot intensities revealed that each compound led to approximately 150-250 gene expression changes that were sustained over at least three adjacent time points. The affected genes could be classified into clusters based on their temporal patterns of differential expression. A common set of 44 genes showed similar expression changes in response to all three compounds. Of these changes, 90% could be confirmed by quantitative RT-PCR analysis. The results indicate that detailed array-based time-course studies, coupled with a sensitive and highly specific confirmation assay, provide a powerful means of identifying cytotoxicity-associated gene expression changes. Electronic Publication