Degradation of a radiolabeled juvenile hormone analog using two insect species

A synthetic insect juvenile hormone analog (a juvenoid), ethylN-[2-[4-[[2,2-(ethylenedioxy)cyclohexyl]methyl]phenox]ethyl]carbamate, which has displayed high biological activity against different insect species and high stability under field conditions, was selected as a biologically active model compound for a study of a juvenile hormone analog degradation. The biologically active compound itself and its three diversely radiolabeled derivatives were applied to the flesh fly (Sarcophaga bullata) or the tsetse fly (Glossina palpalis), respectively. Monitoring of a fate of the applied juvenile hormone analog was carried out using a detection method of the radioactivity microdistribution within the whole insect body in combination with a radio high performance liquid chromatography (radio-HPLC), both of whole-body extracts made in different, but in advance scheduled, time intervals, and of extracts of insect excreta accumulated over an eight-day experiment.     

Modeling individual tree height to diameter ratio for Norway spruce and European beech in Czech Republic

Key message

The purposed spatially explicit and spatially non-explicit height to diameter ratio models can be useful to evaluate the stability of trees and stands for Norway spruce and European beech forests.

Abstract

Height to diameter ratio (HDR) is an individual tree index, also known as slenderness coefficient, and commonly used to evaluate stability of trees and stands. We developed both spatially explicit and spatially non-explicit HDR models for Norway spruce (Picea abies (L.) Karst.) and European beech (Fagus sylvatica L.) using a large dataset collected from fully stem-mapped permanent research plots in various parts of the Czech Republic. Various tree and stand characteristics were evaluated for their potential contributions to the the HDR models. In addition to diameter at breast height (DBH), other highly significant predictor variables identified are dominant height (HDOM) (site quality measure), dominant diameter (DDOM) and quadratic mean diameter (QMD) (spatially non-explicit competition measures), and Hegyi’s index (spatially explicit competition index, CI). A simple exponential decay function was chosen as a base function to include these predictor variables. Both spatially explicit and spatially non-explicit models described large parts of the HDR variations [R _adj ²  = 0.66 (Norway spruce), 0.72 (European beech)] without any systematic deviation of the residuals across the observed data range. Unlike for European beech, spatially explicit model for Norway spruce better described HDR variations than its spatially non-explicit counterpart. After DBH, HDOM provided the largest contribution to each model type, followed by DDOM and QMD or CI for both species. The HDR increased with increasing HDOM and CI, but it decreased with increasing DDOM and QMD, suggesting there were significantly large effects of site quality and stand density on HDR. Because of a little difference between the fit statistics and graphical displays of the two model types, spatially non-explicit model is recommended for prediction of HDR for both species as this model does not require spatially explicit CI, which is computationally much more complex than spatially non-explicit competition measures. The proposed HDR models may be applicable to assess stability of trees and stands, and to regulate stand densities.

     

Mitochondrial pyruvate carrier in Trypanosoma brucei

Pyruvate is a key product of glycolysis that regulates the energy metabolism of cells. In Trypanosoma brucei, the causative agent of sleeping sickness, the fate of pyruvate varies dramatically during the parasite life cycle. In bloodstream forms, pyruvate is mainly excreted, whereas in tsetse fly forms, pyruvate is metabolized in mitochondria yielding additional ATP molecules. The character of the molecular machinery that mediates pyruvate transport across mitochondrial membrane was elusive until the recent discovery of mitochondrial pyruvate carrier (MPC) in yeast and mammals. Here, we characterized pyruvate import into mitochondrion of T. brucei. We identified mpc1 and mpc2 homologs in the T. brucei genome with attributes of MPC protein family and we demonstrated that both proteins are present in the mitochondrial membrane of the parasite. Investigations of mpc1 or mpc2 gene knock‐out cells proved that T. brucei MPC1/2 proteins facilitate mitochondrial pyruvate transport. Interestingly, MPC is expressed not only in procyclic trypanosomes with fully activated mitochondria but also in bloodstream trypanosomes in which most of pyruvate is excreted. Moreover, MPC appears to be essential for bloodstream forms, supporting the recently emerging picture that the functions of mitochondria in bloodstream forms are more diverse than it was originally thought.     

Indigogenic methods for glycosidases

Summary In the histochemical detection of -D-glucosidase the indigogenic method of Pearson et al. was tested and evaluated. 4-Cl-5-Br-3-indolyl--D-glucoside was used as the substrate.Intestinal -D-glucosidase is firmly bound to the structure. About 60% of activity survives 2 hours fixation in cold 4% formaldehyde and some activity can be demonstrated even in paraffin sections after a shortened embedding.The localization obtained with the original method is not correct. Due to a slow oxidation of indoxyl in the acid pH range and to hydrogen peroxide formation indigo is deposited in sites with an active peroxidase or pseudoperoxidase. The addition of horse-radish peroxidase improves the localization but does not entirely prevent diffusion artifacts. A striking improvement of the localization can be achieved by a mixture of ferri-ferrocyanide. 3.1 · 10^–3 M concentration of this oxidation catalyst which is still very effective causes only a 26% inhibition of the enzyme activity as revealed by biochemical assays in homogenates of rat intestine.A new medium was devised consisting of 0.1 M citrate phosphate buffer pH 5.5, 8 · 10^–4 M 4-Cl-5-Br-3-indolyl--D-glucoside and 3.1 · 10^–3 M ferri-ferrocyanide mixture. With this medium a very clear brush border localization of the enzyme activity (activities) in enterocytes of the rat and human intestine was demonstrated. This activity is present in differentiated enterocytes covering the villi. The highest activity resides in the jejunum. Enzyme activity is considerably inhibited by galactonolactone (5 · 10^–3 M) and gluconolactone (4 · 10^–4 M). It is completely inhibited by florizin (5 · 10^–3 M). Cellobiose (8 · 10^–2 M) caused 65%, lactose (8 · 10^–2 M) 48% and glucose (8 · 10^–2 M) 35% inhibition (data were obtained by cytospectrophotometry). In patients with celiac sprue the activity is very much decreased. Its restitution after a gluten-free diet proceeds roughly parallel to that of lactase. The relationship of the demonstrated activity (activities) to florizin hydrolase and lactase is discussed.In the kidney the reaction is very weak and is confined to the cells of proximal convoluted tubules (diffuse staining with some enhancement at the luminal surface of the brush border). The method is also very useful for processing zymograms.     

Synthetic substrates in the histochemical demonstration of intestinal disaccharidases

Summary The splitting of 6-Br-2-naphthyl-, -naphthyl-, and 4-Cl-5-Br-3-indolyl-glycosides which proved useful for the assessment of cytological localization of intestinal enzymes in previous studies was investigated using isolated human and rat intestinal disaccharidases as a source of enzyme activities.Previous findings based on histochemical studies were confirmed and extended. 6-Br-2naphthyl-D-glucoside is cleaved by glucoamylase and sucrase-isomaltase. The participatio of trehalase in splitting of this substrate is very low and can be neglected. The mentioned -glucosidases are responsible for the brush border staining of enterocytes with this substrate when unfixed cold microtome sections are used. Even when a differential heat inactivation of sucrase-isomaltase and of glucoamylase occurs during paraffin embedding (so that the staining in paraffin sections is due mostly to glucoamylase) the use of natural substrates is desirable for a more precise assessment of sucrase-isomaltase activity (but without the possibility of a correct localization).4-Cl-5-Br-3-indolyl--D-fucoside is the substrate of choice for the demonstration of lactase. Even when this substrate is split also by hetero--galactosidase and by acid (lysosomal) -galactosidase these activities do not disturb the histochemical demonstration of lactase. If however some doubts arise, the inhibition with p-Cl-mercuribenzoate (2 · 10^–4 M) is to be emloyed (lactase activity is not inhibited). Due to a low K_m and a high V_max of indolyl-fucoside and due to its extreme stability in solution (which enables to use the substrate solution repeatidly) this substrate is suitable in routine practice even though it is expensive. -naphthyl- and 4-Cl-5-Br-3-indolyl--D-glucosides are split by lactase and -glucosidase. Due to the fact that the mutual delineation of these activities is not easy and that K_m an V_max for lactase are not so favourable as in the case of fucoside these substrates are not recommended for the assessment of lactase.6-Br-2-naphthyl--D-glucoside is the substrate of choice for the histochemical studies concerned with hetero--galactosidase and 4-Cl-5-Br-3-indolyl--D-galactoside for acid -galactosidase.     

Genistella Léger etGauthier vs.Genistella Ortega; a nomenclatural note

The nameLegeriomyces Pouz. is proposed forGenistella Léger etGauthier (Trichomycetes), which is a later homonym ofGenistella Ortega (Papilionaceae). Simultaneously the family nameLegeriomycetaceae Pouz. is proposed to replace the family nameGenistellaceae Léger etGauthier.