Carlos E. Winter

The giant DNA puffs of Rhynchosciara americana code for polypeptides of the salivary gland secretion

Abstract It was found that the polypeptides correlated with the appearance of DNA puffs B2 and C3 in the salivary gland of Rhynchosciara americana are present in the salivary secretion. This makes it very likely that these polypeptides are used in spinning the communal cocoon.

Vitellogenins and other haemolymph proteins involved in the oogenesis of Rhynchosciara americana

Abstract The electrophoretical analysis of haemolymph proteins from Rhynchosciara americana , in the presence of sodium dodecyl sulphate revealed the occurrence of two vitellogenic polypeptides, V 1 and V 2 , with molecular weights of 240,000 and 220,000 respectively. These polypeptides are synthesized by the adult female fat bodies and are found in the yolk granules. The vitellin molecule is formed at least by the polypeptides V 1 , V 2 and two others, V 4 and V 5 with molecular weights of 67,000 and 55,000 respectively. All these polypeptides can be stained by PAS and by methyl green, suggesting that they are phosphoglycopeptides. Haemolymph proteins, other than vitellogenin, participate in egg formation, the most prominent being a monomeric larval protein of molecular weight 47,000. These larval proteins are found in the soluble fraction of the eggs and make a massive contribution to the total egg protein.

Molecular characterization of the C-3 DNA puff gene of Rhynchosciara americana.

We have mapped a region of about 33 kb which includes the transcription unit of the C-3 DNA puff gene of Rhynchosciara americana. The C-3 TU and a region extending approximately 800 bp upstream of the C-3 promoter were characterized. The TU is composed of three exons and produces a 1.1-kb mRNA whose level in salivary glands increases with the expansion of the C-3 puff. The C-3 messenger appears to undergo rapid deadenylation resulting in an RNA of about 0.95 kb which can still be observed in gland cells 15 h after the puff has regressed. The 1.1-kb mRNA codes for a 32.4-kDa, predominantly alpha-helical polypeptide with three conserved parallel coiled-coil stretches. The aa composition and structure of this polypeptide suggests that it is secreted and contributes to the formation of the cocoon in which the larvae pupate. The region upstream of the promoter contains several A-rich sequences with similarity to the ACS of yeast which might have a role in the initiation of replication/amplification.

Underground leaves of Philcoxia trap and digest nematodes

The recently described genus Philcoxia comprises three species restricted to well lit and low-nutrient soils in the Brazilian Cerrado. The morphological and habitat similarities of Philcoxia to those of some carnivorous plants, along with recent observations of nematodes over its subterranean leaves, prompted the suggestion that the genus is carnivorous. Here we report compelling evidence of carnivory in Philcoxia of the Plantaginaceae, a family in which no carnivorous members are otherwise known. We also document both a unique capturing strategy for carnivorous plants and a case of a plant that traps and digests nematodes with underground adhesive leaves. Our findings illustrate how much can still be discovered about the origin, distribution, and frequency of the carnivorous syndrome in angiosperms and, more generally, about the diversity of nutrient-acquisition mechanisms that have evolved in plants growing in severely nutrient-impoverished environments such as the Brazilian Cerrado, one of the worlds 34 biodiversity hotspots.

Effects of Wolbachia on fitness of Culex quinquefasciatus (Diptera; Culicidae).

Wolbachia are α-proteobacteria that were first reported in Culex pipiens mosquitoes early in the twentieth century. Since then, the effect of Wolbachia on their hosts reproduction has drawn attention and has been increasingly investigated. Given the extreme complexity of this interaction, new study cases are welcomed to enhance its understanding. The present work addressed the influence of Wolbachia on Cx. quinquefasciatus, the cosmopolitan member of the Cx. pipiens complex. Samples of a Cx. quinquefasciatus colony (wPip(+)) originated from individuals naturally infected by Wolbachiapipientis B strain, were cured with tetracycline, yielding a Wolbachia-free colony (wPip(-)). Both the presence of bacteria and the efficiency of bacterial elimination were checked by PCR of the wsp gene. Total reproductive unidirectional incompatibility occurred when wPip(-) females were crossed with wPip(+) males, whereas the other three types of reciprocal crosses were viable. Reproductive aspects were also comparatively evaluated between colonies. Concerning oviposition time during the first gonotrophic cycle, wPip(+) females developed and laid eggs earlier than did wPip(-) females. Reproductive fitness was higher among wPip(-) than wPip(+) females regarding the following parameters: fertility: egg rafts/fed females; fecundity: eggs/raft, and viability: larvae/eggs. Conversely, longevity of wPip(-) females was lower. Summarising, although the infected mosquitoes have the advantage of a higher longevity, they have lower reproductive fitness. Our results are partly distinct from all other reports on Aedes and Culex mosquitoes previously published.

Molecular and morphological characterization of heterorhabditid entomopathogenic nematodes from the tropical rainforest in Brazil

Despite massive losses of primary forest, the Amazonian rainforest remains an extremely rich source of biodiversity. In recent years, entomopathogenic nematodes (EPNs) have been isolated from soil in various parts of the world and used successfully as biological control agents against numerous insect pests. Therefore, a sampling in the rainforest of Monte Negro, Rondônia, Brazil was conducted with the aim of discovering new strains and/or species of EPNs for future development as biological control agents. From 156 soil samples taken at nine collecting sites, 19 isolates were obtained, all of them belonging to the genus Heterorhabditis. Four strains were subjected to detailed morphological and molecular evaluation. Based on morphometrics and internal transcribed spacer (ITS) sequence data, the strains LPP1, LPP2 and LPP4 were identified as Heterorhabditis indica, whereas LPP7 was considered Heterorhabditis baujardi. Comparative analysis of the ITS1 sequence of H. indica and H. baujardi isolates showed a polymorphic site for the restriction enzyme Tth 111 that could be used to distinguish the two species. Consequently, strains LPP1, LPP2, LPP3, LPP4, and LPP9 were identified as H. indica, whereas LPP5, LPP7, LPP8 and LPP10 were identified as H. baujardi.

Protein synthesis in the salivary glands of Rhynchosciara americana

Abstract During the fourth larval instar the salivary gland of Rhynchosciara americana is engaged in the synthesis of a silk-like protein secretion. This secretion is composed of a small number of polypeptides whose molecular weights vary between 20,300 and 152,000. The polypeptide composition of this secretion changes as the larvae pass through different periods in the fourth instar toward pupation, and the size of its constituent polypeptides generally decreases. When the larvae stop feeding at the beginning of communal cocoon spinning (third period), an exponential increase in the production of secretion occurs until it represents 75% of the protein synthesis in the gland. In vitro experiments with [ 3 H]leucine incorporation show that all the major secreted polypeptides are produced in the posterior region of the gland, with the anterior region making only a small contribution. Each polypeptide is produced during a specific stage of larval development. Thus the salivary gland follows a precise developmental pattern of protein synthesis which is typical of highly specialized organs.

The yolk polypeptides of a free-living rhabditid nematode

Abstract 1. 1. The yolk proteins of hermaphrodite Dolichorhabditis sp. (Nematode, Rhabditida) are composed of at least three polypeptides: VT1, VT2 and VT3 with molecular masses of 175.2, 107 and 82 kDa respectively. 2. 2. All three yolk polypeptides make up at least one native protein complex which can be resolved by PAGE. 3. 3. The yolk proteins are glycosylated and can be isolated by chromatography in Con A-Sepharose. 4. 4. Partial chymotryptic hydrolysis shows that VT2 in different from its C. elegans homologue, YP115. 5. 5. The main polypeptides synthesized by whole animals are the yolk components which are actively secreted in the incubation medium.

CHARACTERIZATION OF THE MAIN PLASMA LIPOPROTEINS FROM THE OVOVIVIPAROUS VIPERID SNAKE BOTHROPS JARARACA

Abstract Plasma from estrogen treated males of Bothropsjararaca fractionated by ultracentrifugation showed three main fractions with very low density, intermediate density and high density, respectively. The very low density and high density fractions are mainly composed of lipoproteins detected by specific staining with Sudan Black. Electron microscopy of the very low density lipoproteins shows a single population of particles with a mean diameter of 29.6 nm similar to that of very low density lipoproteins from laying hens. The very low density lipoprotein of Bothrops jararaca is composed of two polypeptides with molecular masses of 450 and 20 kDa. Three main polypeptides are detected in the plasma high density fraction with molecular masses of 190, 160 and 110 kDa. Analysis of whole plasma proteins, from control and estrogen treated males, by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the 20 kDa very low density lipoprotein polypeptide and the 160 and 110 kDa high density lipoprotein polypeptides are induced de novo after estrogen treatment. These results suggest that the two very low density lipoprotein polypeptides are the snake counterparts of chicken very low density polypepudes. The estrogen induced high density lipoprotein polypeptides probably make up the vitellogenin(s) of Bothropsjararaca . An antiserum raised against the whole very low density plasma fraction was able to recognize only the two polypepudes found in this fraction in the plasma of treated males. Using this anuserum, in a double immunodiffusion test, we were able to detect only a faint reaction against Bothrops jararaca egg yolk proteins.

Vitellin-binding proteins in the nematode Oscheius tipulae (Nematoda, Rhabditida)

We describe the first application of a non-radioactive ligand-blotting technique to the characterization of proteins interacting with nematode vitellins. Chromatographically purified vitellins from the free-living nematode Oscheius tipulae were labeled with fluorescein in vitro. Ligand-blotting assays with horseradish peroxidase-conjugated anti-fluorescein antibodies showed that labeled vitellins reacted specifically with a polypeptide of approximately 100 kDa, which we named P100. This polypeptide is a specific worms vitellin-binding protein that is present only in adult worms. Blots containing purified O. tipulae vitellin preparations showed no detectable signal in the 100 kDa region, ruling out any possibility of yolk polypeptides self-assembling under the conditions used in our assay. Experiments done in the presence of alpha-methyl mannoside ruled out the possibility of vitellins binding to P100 through mannose residues. Triton X-114 fractionation of whole worm extracts showed that P100 is either a membrane protein or has highly hydrophobic regions.