Purification and characterization of glycosyltransferases involved in anthocyanin biosynthesis in cell-suspension cultures of Daucus carota L.

The major anthocyanins accumulated by an Afghan cultivar of Daucus carota L. are cyanidin 3-(xylosylglucosylgalactosides) acylated with sinapic or ferulic acid. The formation of the branched triglycoside present as a common structural element requires an ordered sequence of glycosylation events. Two of these enzymic glycosylation reactions have been detected in protein preparations from carrot cell-suspension cultures. The first step is a galactosyl transfer catalyzed by UDP-galactose: cyanidin galactosyltransferase (CGT) resulting in cyanidin 3-galactoside. The putative second step is the formation of cyanidin 3-(xylosylgalactoside) catalyzed by UDP-xylose: cyanidin 3-galactoside xylosyltransferase (CGXT). Both enzyme activities were characterized from crude protein preparations. The CGT was purified 526-fold from the cytosolic fraction of UV-irradiated cell cultures by ion-exchange chromatography on diethylaminoethyl (DEAE)-Sephacel, affinity chromatography on Blue Sepharose CL-6B, gel permeation chromatography on Sephadex G-75 and elution from the gel matrix after non-dissociating PAGE. Its molecular mass was estimated by SDS-PAGE and by calibrated gel permeation chromatography on Sephadex G-75. In both cases a molecular mass of 52 kDa was determined, indicating that the native protein is a monomer of 52 kDa. The galactosyl transfer and the xylosyl transfer are presumed to be catalyzed by separate enzymes.

[Clinical and epidemiologic aspects of Aujeszky's disease in sheep]. / Klinische und epidemiologische Aspekte der Aujeszky-Krankheit des Schafes.

Aujeszky's disease virus is spread by horizontal transmission from infected pigs to sheep with high losses. Clinical symptoms in sheep, for example pruritus, are obvious. Infection of sheep can be avoided by strictly separating pigs and sheep. Vaccination of pigs therefore indirectly also gives good protection to other susceptible species on the farm.

Serological diagnosis of Aujeszky's disease: a comparison between reactions of blood samples from sows and their offspring.

Blood sera from sows and their piglets were compared for their suitability for the serological diagnosis of Aujeszky's disease. Within a few days after parturition, blood and colostrum samples were collected from a total of 104 sows from 8 different gI-vaccinated breeding herds. Three piglets of each litter were bled simultaneously with their mother and again 3 weeks later. All 416 sera reacted positively in the screening for vaccination-induced antibodies. Using the gI ELISA, the sera of 16 sows and their offspring reacted positively, while 86 sows and their piglets reacted negatively and 2 sows and their piglets showed reactions near the test-cutoff. By testing piglets from another 1,300 sows in 37 farms, the practicability of the proposed method was proved. Blood samples were taken at intervals of 4 to 6 weeks from piglets of all sows, which had farrowed in the meantime. Within a six to nine months period, the serological status of a breeding herd could be established.

Constancy of Aujeszky's field virus antibody titres in sows repeatedly vaccinated with a gI-negative vaccine.

A breeding herd repeatedly vaccinated with a gI-deleted vaccine was regularly tested serologically for Aujeszky's field virus antibodies over a period of up to 35 months. Of the 100 sows tested, 18 animals reacted positively with constant titers during the whole period of the investigation. On the other hand, there was no serological evidence that any of the contact animals contracted field virus infection.

Continuous-risk utility assessment in medical decision making.

The authors argue that for risky medical treatment decisions, conventional utility assessment techniques are inadequate due to their emphasis on unrealistic risk magnitudes and sure consequences, leading to assessment questions that are unfamiliar to most patients, have little educational value, and do not reliably extend to the application at hand. As an alternative, medical utility assessments should be performed in a continuous-risk domain with risk levels matching those of the actual decision problem. In support of this position, the authors describe an empirical study that compares the responses of subjects under a conventional assessment protocol with those of subjects under a continuous-risk utility assessment protocol. Preliminary results show that conventional assessment protocols result in significantly lower estimates of the degrees of risk aversion compared with a more realistic continuous-risk protocol.

[Induction of humoral antibodies by a BHV-1 intradermal test]. / Induktion humoraler Antikörper durch einen BHV 1-Intrakutantest.

An intradermal test for the diagnosis of BHV 1 (Intrakutantest Behringwerke AG) was applied to 53 nonvaccinated BHV 1-seronegative cattle aged 7 months to 8 years. Serologic blood testing performed subsequently using ELISA (Enzygnost-IBR/IPV, Behringwerke AG) and SNT revealed seroconversion in 24 of 45 animals without previous maternal antibodies. A second application of the intradermal test after these BHV 1-antibodies had declined, lead to a 'booster-effect' while a control group remained negative. Six of nine animals not affected by the first intradermal application of BHV 1-antigen, developed BHV 1-antibodies following the second intradermal test. Eight animals possessing maternal antibodies showed no serological response to the intradermal test at all. Present results strongly suggest an induction of humoral BHV 1-antibodies by the intradermal application of inactivated BHV 1. Consequently, the indirect control of BHV 1 infections, especially in cattle breeding farms, should be done exclusively by serological examination.

The uptake of acylated anthocyanin into isolated vacuoles from a cell suspension culture of Daucus carota.

Anthocyanin-containing vacuoles were isolated from protoplasts of a cell suspension culture of Daucus carota. The vacuoles were stable for at least 2 h as demonstrated by the fact that they showed no efflux of anthocyanin. The uptake of radioactively labelled anthocyanin was time-dependent with a pH optimum at 7.5, and could be inhibited by the protonophore carbonylcyanide m-chlorophenylhydrazone. Furthermore, the transport was specific, since vacuoles from other plant species showed no uptake of labelled anthocyanin, and strongly depended on acylation with sinapic acid, as deacylated glycosides were not taken up by isolated vacuoles. Hence, it is suggested that the acylation of anthocyanin, which is also required for the stabilization of colour in vacuoles, is important for transport, and that acylated anthocyanin is transported by a selective carrier and might be trapped by a pH-dependent conformational change of the molecule inside the acid vacuolar sap.