Biological studies on Trichogrammatoidea bactrae Nagaraja (Hymenoptera: Trichogrammatidae), egg parasitoid of Tuta absoluta Meyrick (Lepidoptera: Gelechiidae)

The tomato moth, Tuta absoluta Meyrick, is one of the most important tomato pests in South America. In Argentina, management strategies include only chemical control. In this work, the parasitoid wasp Trichogrammatoidea bactrae Nagaraja was evaluated as a potential natural enemy against this pest. Biological and population parameters were estimated by developing a life table under laboratory conditions at 25 ± 1oC, 14:10 photoperiod and 60 ± 10% RH. Three cohorts of 26-30 T. bactrae females each were placed with one of the three following treatments: 1 - Sitotroga cerealella (Olivier) eggs on a piece of cardboard; 2 - S. cerealella eggs on a piece of tomato leaf, and 3- T. absoluta eggs on a piece on tomato leaf. The following parameters were estimated for each cohort: survival (egg to adult), longevity, fecundity and oviposition period of females, sex proportion of the F1, net rate of reproduction (Ro), mean generation time (T) and intrinsic rate of population increase (r m). Survival of the T. bactrae immatures was higher than 90% on both, S. cerealella and T. absoluta eggs. The female survival curves corresponded to type III and showed no significant differences among treatments. The three cohorts did not show significant differences between sex ratio, female longevity, oviposition period, fecundity and the population parameters studied. These results indicate that T. bactrae would be a potential biological control agent of T. absoluta.     

Annexin 6 modulates the maxi-chloride channel of the apical membrane of syncytiotrophoblast isolated from human placenta

The syncytiotrophoblast separates the maternal and fetal blood and constitutes the primary barrier for maternal-fetal transport. The Maxi-chloride channel from the apical membrane of the syncytiotrophoblast plays a role in the chloride conductance. Annexins can play an important role in the regulation of membrane events. In this study we evaluate the role of annexin 6 in the Maxichloride channel properties. The results showed that annexin 6 is bound in the apical placenta membranes in a calcium-dependent phospholipid-binding manner but also in a calcium-independent fashion. The neutralization of annexin 6 decreased the total current by 39 +/- 1.9% in the range of +/-80 mV, and the currents decrease with the time. The single-channel slope conductance was decreased from 253 +/- 7.4 pS (control) to 105 +/- 13 pS, and the amplitude decreased by 50%. The open probability was also affected when higher voltage steps were used, changes in either the positive or negative direction induced the channel to close, and the open probability (P(o)) did not decrease. In channels with neutralized annexin 6, it was maintained at 1 at +/-40 mV and at +/-80 mV. These results suggest that endogenous annexin 6 could regulate the Maxi-chloride channel. The results obtained with normal placentae, in which annexin 6 was neutralized, are similar to those described for the Maxi-chloride channel isolated from pre-eclamptic placenta. Together these data suggest that annexin 6 could play an important role in ion transport of the placenta.     

Calcium waves propagate through radial glial cells and modulate proliferation in the developing neocortex

The majority of neurons in the adult neocortex are produced embryonically during a brief but intense period of neuronal proliferation. The radial glial cell, a transient embryonic cell type known for its crucial role in neuronal migration, has recently been shown to function as a neuronal progenitor cell and appears to produce most cortical pyramidal neurons. Radial glial cell modulation could thus affect neuron production, neuronal migration, and overall cortical architecture; however, signaling mechanisms among radial glia have not been studied directly. We demonstrate here that calcium waves propagate through radial glial cells in the proliferative cortical ventricular zone (VZ). Radial glial calcium waves occur spontaneously and require connexin hemichannels, P2Y1 ATP receptors, and intracellular IP3-mediated calcium release. Furthermore, we show that wave disruption decreases VZ proliferation during the peak of embryonic neurogenesis. Taken together, these results demonstrate a radial glial signaling mechanism that may regulate cortical neuronal production.     

Efficient GFP expression in the mushrooms Agaricus bisporus and Coprinus cinereus requires introns

We have developed a "Molecular Toolkit" comprising interchangeable promoters and marker genes to facilitate transformation of homobasidiomycete mushrooms. We describe the evaluation of a range of promoters in the homobasidiomycetes Agaricus bisporus and Coprinus cinereus using green fluorescent protein (GFP) as a reporter gene; the C. cinereus trp1 promoter and A. bisporus trp2 and gpdII promoters proving successful in driving expression in C. cinereus, with the gpdII promoter also functioning in A. bisporus. Our investigations demonstrate that a prerequisite for GFP expression in C. cinereus and A. bisporus is the presence of an intron. This is the first reported expression of GFP in either C. cinereus or A. bisporus.     

The effects of ropy-1 mutation on cytoplasmic organization and intracellular motility in mature hyphae of Neurospora crassa

We have used light and electron microscopy to document the cytoplasmic effects of the ropy (ro-1) mutation in mature hyphae of Neurospora crassa and to better understand the role(s) of dynein during hyphal tip growth. Based on video-enhanced DIC light microscopy, the mature, growing hyphae of N. crassa wild type could be divided into four regions according to cytoplasmic organization and behavior: the apical region (I) and three subapical regions (II, III, and IV). A well-defined Spitzenk?rper dominated the cytoplasm of region I. In region II, vesicles ( approximately 0.48 micro m diameter) and mitochondria maintained primarily a constant location within the advancing cytoplasm. This region was typically void of nuclei. Vesicles exhibited anterograde and retrograde motility in regions III and IV and followed generally parallel paths along the longitudinal axis of the cell. A small population of mitochondria displayed rapid anterograde and retrograde movements, while most maintained a constant position in the advancing cytoplasm in regions III and IV. Many nuclei occupied the cytoplasm of regions III and IV. In ro-1 hyphae, discrete cytoplasmic regions were not recognized and the motility and/or positioning of vesicles, mitochondria, and nuclei were altered to varying degrees, relative to the wild type cells. Immunofluorescence microscopy revealed that the microtubule cytoskeleton was severely disrupted in ro-1 cells. Transmission electron microscopy of cryofixed cells confirmed that region I of wild-type hyphae contained a Spitzenk?rper composed of an aggregation of small apical vesicles that surrounded entirely or partially a central core composed, in part, of microvesicles embedded in a dense granular to fibrillar matrix. The apex of ro-1 the hypha contained a Spitzenk?rper with reduced numbers of apical vesicles but maintained a defined central core. Clearly, dynein deficiency in the mutant caused profound perturbation in microtubule organization and function and, consequently, organelle dynamics and positioning. These perturbations impact negatively on the organization and stability of the Spitzenk?rper, which, in turn, led to severe reduction in growth rate and altered hyphal morphology.     

Complex viscoelasticity of normal and lectin treated erythrocytes using laser diffractometry.

A new method to find directly complex viscoelastic parameters (CVP) of red blood cells (RBC) is presented in this paper. Experimental determinations were carried out in an Erythrodeformeter (Rasia et al., 1986) operating in oscillating mode (0.5 to 3.5 Hz). The Erythrodeformeter performs direct determination of CVP of erythrocytes undergoing sinusoidal shear stresses by laser diffractometry. The measurements lead to the determination of mean values of the four components of erythrocyte complex viscoelasticity. The influence of the alterations induced on erythrocyte membrane by vegetable lactins (Ulex europaeus, wheat germ agglutinin and Enterolobium contorticilicum seeds) was analyzed to verify the sensitivity of this method. Differences observed between the CVP parameters of treated cells and the ones corresponding to control samples (non treated cells) are analyzed. Results obtained from cells treated with wheat germ agglutinin agree with observations published by Smith and Hochmuth (1982). Determinations of RBC complex viscoelasticity carried out by laser diffractometry could become an important tool to understand the influence of the factors associated with alterations of the rheologic properties of RBC membrane, which can affect the in vivo blood flow.     

Characterization of a Chlorophyta microalga isolated from a microbial mat in Salar de Atacama (northern Chile) as a potential source of compounds for biotechnological applications

Microalgae are an important source of unsaturated fatty acids, phospholipids, glycolipids, and carotenes, which are useful compounds for the food and pharmaceutical industries. The Atacama Desert of northern Chile is one of the driest deserts on Earth and, as such, it is a great natural laboratory in which to study new microorganisms adapted to extreme environments. A microalgal strain, referred to here as CH03, was isolated from a microbial mat in salt flat water in Salar de Atacama. Genetic analysis of the 18S ribosomal RNA gene showed that the strain had homology with other known sequences of the species Chlorella sorokiniana. Our results revealed the adaptability of this microalga to freshwater medium under laboratory conditions, despite coming from an extremely high‐salinity environment. The fatty acid profile of CH03(A) newly isolated in Bold's basal medium differed from that of CH03(B) cultured in vitro in modified F/2 medium and from another five strains of C. sorokiniana and three strains of Chlorella vulgaris in that it had a high stearic acid content and had no polyunsaturated fatty acids. The major biochemical components observed in this strain were proteins (64.3–73.6%) and lipids (26.6–32.6%). This study suggests that the strain CH03 could be a protein source and that this oleaginous microalga is easy to grow in vitro as a biological model for future studies.     

Outdoor pilot-scale production of Botryococcus braunii in panel reactors

In this paper, the outdoor production of Botryococcus braunii in pilot-scale panel reactors (0.4?m³) is studied under uncontrolled conditions at a location close to the Atacama Desert (Chile). Discontinuous experiments were performed on different dates to determine the feasibility of the culture and the influence of environmental conditions on the system yield. Data showed that solar radiation is a major parameter in determining system yield, the average irradiance inside the culture determining both the growth rate and biomass productivity. A maximum specific growth rate of 0.09?day^?1 and biomass productivity of 0.02?g?L^?1?day^?1 (dry weight) were measured in discontinuous mode, at an average irradiance of 60?μE?m^?2?s^?1. With respect to lipids, a productivity of 2.5?mg?L^?1?day^?1 was obtained under favourable growth conditions; no accumulation of lipids at the stationary phase was observed. To confirm this behaviour, a semicontinuous culture was performed at 0.04?day^?1 in a larger reactor (1?m³). In this experiment, the biomass concentration and productivity was 0.3?g?L^?1 and 0.015?g?L^?1?day^?1, respectively. The lipid content and productivity was 15.6% and 2.4?mg?L^?1?day^?1, respectively, the mean average irradiance inside the reactor being 60?μmol photons?m^?2?s^?1. The light path of the reactor determines the light availability, thus determining also the biomass concentration and productivity of the reactor once the dilution rate is fixed. Experimentally, biomass productivity of 0.015?g?L^?1?day^?1 was determined for a light path of 0.15?m, but this can be increased by more than three times for a light path of 0.1?m. These data confirm that this alga can be produced outdoors in a secure form, the culture yield improving when optimal conditions are applied, the data reported here establishing the starting point for the development of the process.     

Influence of C-terminal protein domains and protein-lipid interactions on tetramerization and stability of the potassium channel KcsA

KcsA is a prokaryotic potassium channel formed by the assembly of four identical subunits around a central aqueous pore. Although the high-resolution X-ray structure of the transmembrane portion of KcsA is known [Doyle, D. A., Morais, C. J., Pfuetzner, R. A., Kuo, A., Gulbis, J. M., Cohen, S. L., Chait, B. T., and MacKinnon, R. (1998) Science 280, 69-77], the identification of the molecular determinant(s) involved in promoting subunit tetramerization remains to be determined. Here, C-terminal deletion channel mutants, KcsA Delta125-160 and Delta120-160, as well as 1-125 KcsA obtained from chymotrypsin cleavage of full-length 1-160 KcsA, have been used to evaluate the role of the C-terminal segment on the stability and tetrameric assembly of the channel protein. We found that the lack of the cytoplasmic C-terminal domain of KcsA, and most critically the 120-124 sequence stretch, impairs tetrameric assembly of channel subunits in a heterologous E. coli expression system. Molecular modeling of KcsA predicts that, indeed, such sequence stretch provides intersubunit interaction sites by hydrogen bonding to amino acid residues in N- and C-terminal segments of adjacent subunits. However, once the KcsA tetramer is assembled, its remarkable in vitro stability to detergent or to heat-induced dissociation into subunits is not greatly influenced by whether the entire C-terminal domain continues being part of the protein. Finally and most interestingly, it is observed that, even in the absence of the C-terminal domain involved in tetramerization, reconstitution into membrane lipids promotes in vitro KcsA tetramerization very efficiently, an event which is likely mediated by allowing proper hydrophobic interactions involving intramembrane protein domains.