Trypanosoma brucei: morphometric changes and loss of infectivity during transformation of bloodstream forms to procyclic culture forms in vitro.

The transformation of Trypanosoma brucei bloodstream forms to procyclic culture forms in the semidefined medium SDM-77 has been studied by light microscopy and quantitative electron microscopy. Stumpy and intermediate forms are able to transform to culture forms whereas slender forms die after approximately 24 hr. The surface coat and infectivity for the mammalian host are lost after 72 hr. Morphometrical analysis of the cells during transformation process revealed: (1) The cytoplasm and the cell surface increased significantly; (2) the volume density of the mitochondrion increased twofold and the surface density of the inner mitochondrial membrane increased threefold; (3) the volume density of the glycosomes remained about constant; (4) the volume density of the lipid inclusions increased up to 72 hr, probably as a result of the complete oxidation of glucose. Transformation as observed by light microscopy was completed in 72 hr. However, quantitative electron microscopy revealed that establishment of the culture form was incomplete even after 11 days.     

Iridoid glucosides from Melampyrum

Melampyrum arvense and M. cristatum contain, besides aucubin, 8-epiloganin and melampyroside, a new natural iridoid glucoside: gardoside methyl ester. In addition, M. arvense contains mussaenoside and M. cristatum mussaenosidic acid, another novel iridoid glucoside.     

Early cerebral metabolic and electrophysiological recovery during controlled hypoxemic resuscitation in piglets

We tested the hypothesis that controlledhypoxemic resuscitation improves early cerebral metabolic andelectrophysiological recovery in hypoxic newborn piglets. Severelyhypoxic anesthetized piglets were randomly divided into threeresuscitation groups: hypoxemic, 21%O₂, and 100%O₂ groups (8 in each group). Thehypoxemic group was mechanically ventilated with 12-18%O₂ adjusted to achieve a cerebralvenous O₂ saturation of17-23% (baseline; 45 ± 1%). Base excess (BE) reached22 ± 1 mM at the end of hypoxia. During a 2-h resuscitationperiod, no significant differences in time to recovery ofelectroencephalography (EEG), quality of EEG at recovery, orextracellular hypoxanthine concentrations in the cerebral cortex andstriatum were found among the groups. BE and plasma hypoxanthine,however, normalized significantly more slowly during controlledhypoxemic resuscitation than during resuscitation with 21 or 100%O₂. We conclude that early brainrecovery during controlled hypoxemic resuscitation was as efficient as,but not superior to, recovery during resuscitation with 21 or 100%O₂. The systemic metabolicrecovery from hypoxia, however, was delayed during controlled hypoxemicresuscitation.

     

Results of the seventh joint pesticide testing programme carried out by the IOBC/WPRS-Working Group ‘Pesticides and Beneficial Organisms’

The side effect of 10 insecticides, 5 fungicides and 5 herbicides on 24 different species of beneficial organisms was tested by members of the Working Group Pesticides and Beneficial Organisms of the International Organization for Biological Control (IOBC), West Palaearctic Regional Section (WPRS). The tests were conducted by 32 members in 12 countries according to internationally approved guidelines.The microbial insecticides Bacillus thuringiensis var. kurstaki (Delfin), B. thuringiensis var. tenebrionis (Novodor) and Verticillium lecanii (Micro Germin), the fungicides cyproconazol (Alto), difenoconazol (Score), lecithin (Bioblatt Mehltau) and penconazol (Omnex), and the herbicides ethofumesat (Tramat), fluroxypyr (Starane), haloxyfop (Gallant), isoproturon (Arelon) and metamitron (Goltix) were harmless to nearly all the beneficial arthropods. The benzoylurea's teflubenzuron (Nomolt) and flufenoxuron (Cascade) affected predators such as anthocorids, earwigs, coccinellids and lacewings. The remaining preparations were more toxic and should therefore be further tested in semi-field and field experiments on relevant organisms. Most tested fungicides were toxic for the entomopathogenic fungi.     

Exogenous glutamine requirement is confined to late events of T cell activation

Glutamine is required for the proliferation of lymphocytes, but quantitative effects on discrete steps of activation remain unknown to date. Therefore the influence of glutamine (range: 0 mM–1 mM) on the in vitro response of human peripheral blood mononuclear cells (PBMC) to a mitogenic anti-CD3 monoclonal antibody (mAb) was investigated. Expression of surface activation markers by flow cytometry, presence of mRNA of cytokine genes by polymerase chain reaction, release of cytokines by ELISA, and entering into the cell cycle by flow cytometry were sequentially analyzed. Proliferation was measured by a ³H-thymidine incorporation assay. mRNA coding for IL-2, IL-2 receptor, IL-4, IL-5, GM-CSF, and IFN-γ was detectable independently from exogenous glutamine provision; expression of the cell surface activation marker CD69 was also glutamine independent. In contrast, later activation events including the expression of the surface activation markers CD25, CD45RO, and CD71 as well as the production of IFN-γ were found to require exogenous glutamine supply. In contrast, production of TNF-α could be observed in the absence of glutamine and was increased to a limited extent by exogenous glutamine. The overall lymphocyte response as reflected by entering into the cell cycle and proliferation was directly correlated with the glutamine concentration of the culture medium. Efficient progression through the cell cycle was found to require at least 0.5 mM glutamine and an increase in glutamine concentration from 0.1 mM to 1 mM enhanced proliferation by 50%. These results were supported by data obtained following anti-CD3 stimulation of a CD4⁺ T cell clone. Altogether, these data underline that a complete cellular immune response depends on an exogenous glutamine supply. Regarding glutamine requirements, they define early, glutamine-independent and late, glutamine-dependent lymphocyte activation stages.     

Studies of an 800-kilobase DNA stretch of the Drosophila X chromosome: comapping of a subclass of scaffold-attached regions with sequences able to replicate autonomously in Saccharomyces cerevisiae.

We have previously mapped scaffold-attached regions (SARs) on an 800-kilobase DNA walk from the Drosophila X chromosome. We have also previously shown that the strength of binding, i.e., the ability of SARs to bind to all nuclear scaffolds or only to a fraction of them varied from one SAR to another one. In the present study, 71 of the 85 subfragments that bind scaffolds and 38 fragments that do not bind scaffolds were tested for their ability to promote autonomous replicating sequence (ARS) activity in Saccharomyces cerevisiae. Sixteen SAR-containing fragments from the chromosome walk were also examined for association to yeast nuclear scaffolds in vitro. All identified ARSs (a total of 27) were present on SAR-containing fragments, except two, which were adjacent to SARs. There is thus a correlation between ARS and SAR activities, and this correlation defines a SAR subclass. Moreover, the presence of an ARS on a DNA fragment appeared to be highly correlated with the strength of binding. The binding activity was highly conserved from Drosophila melanogaster to yeast. These data suggest that Drosophila DNA sequences responsible for binding to components of the nuclear scaffold from either D. melanogaster or yeast may be involved in the process of heterologous extrachromosomal replication in yeasts.     

Oviposition behavior of the cabbage root fly,Delia radicum (L.), influenced by host plant extracts

An ethanolic extract of cabbage leaves (Brassica oleraceavar. capitata,Golden Acre)and derived fractions were tested on natural and surrogate leaves in order to study the oviposition behavior of the cabbage root fly Delia radicum(Diptera: Anthomyiidae). On surrogate plastic leaves coated with a thin layer of paraffin wax and treated with 0.1 gram leaf equivalent (gle) of an ethanolic raw cabbage extract, the females displayed the same sequence of behavioral patterns as on a natural host plant. The quantified oviposition behavior correlated well with the actual number of eggs laid. The extract-treated surrogate leaves received at least as many eggs as natural leaves with a similar surface area. Previous exposure to surrogate or natural leaves seemed not to induce a specific preference. The three fractions (hexane, butanol, and water) of the raw extract stimulated the oviposition behavior. This was taken as evidence for the presence of a multicomponent mixture or a chemical pattern in the cabbage leaves stimulating oviposition. At the tested concentration each fraction alone could stimulate in some individuals the complete behavior. No effect of volatile components of the raw extract could be detected. This is thought to be due to the extraction procedure, which limits the production of attractive, volatile compounds, such as isothiocyanates.     

A bioassay using the measurement of the growth inhibition of a ciliate protozoan: Colpidium campylum Stokes

A bioassay method using the ciliate protozoan Colpidium campylum is presented in a standardized form. The influence of the initial cell concentration on the potassium dichromate EC50 values was determined. Two intercalibration experiments between two laboratories were performed on ten toxicants in two different conditions. The potassium dichromate EC50 determinations performed by eight different people are also presented. All results are discussed in terms of feasibility and reproducibility of the method, fields of application, and limitations.     

Inhibitors of adenosine consuming parasites through polymer-assisted solution phase synthesis of lipophilic 5′-amido-5′-deoxyadenosine derivatives

Given the more or less global spread of multidrug-resistant plasmodia, structurally diverse starting points for the development of chemotherapeutic agents for the treatment of malaria are urgently needed. Thus, a series of 20 adenosine derivatives with a large lipophilic substituent in N⁶-position were prepared in order to evaluate their potential to inhibit the chloroquine resistant Plasmodium falciparum strain K1 in vitro. The rationale for synthesis of these structures was the high probability of interactions with multiple adenosine associated targets and the assumption that a large hydrophobic N⁶-(4-phenoxy)benzyl substitution should allow the molecules to diffuse across parasite membranes. Starting from readily available inosine, the new compounds were prepared as single isomers using a polymer-assisted acylation protocol enabling the straightforward isolation of the target compounds in pure form. Heterocyclic ring systems were synthesized on-bead on Kenner’s safety-catch linker prior to acylation of the scaffold in solution. Most of the highly pure compounds displayed anti-plasmodial activity in the low micromolar or even submicromolar concentration range.