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At least 973 records · Page 54Linked to original sources
PubMed1993

Nitric oxide synthase activity from a hematophagous insect salivary gland.

The salivary glands of the hematophagous insect, Rhodnius prolixus, contain a nitrosylhemeprotein that dissociates its ligand, NO, to the host tissues while the insect is searching for a blood meal. We now report a salivary nitric oxide synthase activity in this insect. The activity is dependent on NADPH, FAD, tetrahydrobiopterin, calmodulin, Ca2+, and converts arginine to citrulline while producing vasorelaxing activity. Molecular sieving indicates a molecular weight of 185 kDa, coeluting with a diaphorase activity. Results indicate similarity of this insect activity to the vertebrate constitutive NO synthase, suggesting NO synthesis is an evolutionary old biological pathway.

Amino Acid Oxidoreductases↗
PubMed1996

Highly efficient production of GFP and its derivatives in insect cells for visual in vitro applications.

We have generated recombinant baculoviruses for expression of the green fluorescent protein (GFP), a bright GFP mutant (S65T), and a GFP-streptavidin fusion protein in Sf9 and High Five insect cell lines. At 3-4 days post infection, about 30% of the total protein contents was represented by the recombinant protein products, giving the infected insect cells a bright green color which was clearly visible by eye in daylight. The isolated GFP-streptavidin fusion protein, which possessed fluorescence properties identical to those of the native GFP, was capable of binding biotin as shown by using biotinylated beads as well as biotinylated antibody complexes decorating surface expressed GluR-6 glutamate receptor in live and fixed insect cells. The exceptionally high expression levels of GFP and GFP (S65T) and the GFP-streptavidin fusion protein in recombinant baculovirus infected insects should facilitate production of GFP derivatives for in vitro applications.

Animals↗
PubMed1994

Amino acid racemization in amber-entombed insects: implications for DNA preservation.

DNA depurination and amino acid racemization take place at similar rates in aqueous solution at neutral pH. This relationship suggests that amino acid racemization may be useful in accessing the extent of DNA chain breakage in ancient biological remains. To test this suggestion, we have investigated the amino acids in insects entombed in fossilized tree resins ranging in age from <100 years to 130 million years. The amino acids present in 40 to 130 million year old amber-entombed insects resemble those in a modern fly and are probably the most ancient, unaltered amino acids found so far on Earth. In comparison to other geochemical environments on the surface of the Earth, the amino acid racemization rate in amber insect inclusions is retarded by a factor of >10(4). These results suggest that in amber insect inclusions DNA depurination rates would also likely be retarded in comparison to aqueous solution measurements, and thus DNA fragments containing many hundreds of base pairs should be preserved. This conclusion is consistent with the reported successful retrieval of DNA sequences from amber-entombed organisms.

Amber↗
PubMed1994

Digestive enzymes, haemolysins and symbionts in the search for vaccines against blood-sucking insects.

Control of blood-sucking insects by vaccination of the host is an appealing possibility. In this overview the potential of digestive enzymes, haemolysins and symbionts as targets for vaccines are discussed. Blood-sucking insects are largely dependent on proteases for digestion. This is advantageous in the search for a vaccine as there is a considerable fund of biochemical knowledge for this group of molecules. Also there are a range of inhibitors available which can be used to mimic the effects of a vaccine providing a useful guide to the potential usefulness of such a vaccine before the considerable investment needed in producing one. Weighing against this, proteolytic enzymes by their nature will attack antibodies approaching them making it questionable if antibodies ingested by the insect could succeed before they themselves are destroyed by their target. Proteolytic enzymes are also poor immunogens, probably because they attack the antibody making machinery approaching them and because the complexes formed between the abundant vertebrate antiproteolytic molecules and proteases provide a different antigenic target to the native enzymes. Despite their relative paucity, evidence suggests that lipid digesting enzymes deserve attention as potential vaccine targets. Because about 80% of the nutrients in blood are locked up in its formed elements haemolysins are a strong potential target of a vaccine. The obligate haematophages rely on symbionts to provide essential nutrients and in consequence these are also potential vaccine targets in these insects.

Animals↗
PubMed1991

Retarded processing of influenza virus hemagglutinin in insect cells.

When expressed in Spodoptera frugiperda cells by a baculovirus vector, the hemagglutinin of fowl plague virus has been found to contain palmitic acid in covalent hydroxylamine-sensitive linkage, indicating that these cells have the capacity to acylate foreign proteins at cysteine residues. Centrifugation on sucrose density gradients and immune precipitation with conformation-specific antibodies were used to compare trimerization of the hemagglutinin in insect cells and in fowl plague virus-infected MDCK cells. Trimerization of the hemagglutinin was incomplete in insect cells, and the kinetics of this reaction were about three times slower than in vertebrate cells. Similarly, post-translational proteolytic cleavage occurred in insect cells with a half-time of 90 min, and a substantial fraction of the hemagglutinin persisted in uncleaved form. In contrast, hemagglutinin was almost completely cleaved in MDCK cells, and the half-time of cleavage was only 30 min. The data indicate that in insect cells trimerization and, as a result, the subsequent processing steps of the hemagglutinin, are retarded and less efficient. The possible roles of aberrant glycosylation, acidic milieu, and lack of other influenza virus proteins in hemagglutinin trimerization are discussed.

Acylation↗
PubMed1992

Expression of bovine immunodeficiency-like virus envelope glycoproteins by a recombinant baculovirus in insect cells.

The bovine immunodeficiency-like virus (BIV) env open reading frame (ORF) contains both sequences encoding env and sequences for exon 1 of the putative rev gene. Recombinant baculoviruses incorporating BIV env ORF sequences were constructed to characterize the expression, processing, and immunogenicity of products of the BIV env ORF in insect cells and to develop reagents to study native BIV Env glycoproteins. A recombinant baculovirus containing the entire env ORF synthesized a nonglycosylated, 20-kDa, BIV-specific protein, apparently unrelated to native BIV Env proteins. In contrast, a recombinant baculovirus containing a truncated env ORF in which the coding sequences for rev exon 1 were deleted synthesized three size classes of glycosylated proteins in insect cells related to the BIV Env precursor (gp145), surface (gp100), and transmembrane (gp45) glycoproteins observed in BIV-infected mammalian cells. Oligomers of recombinant BIV Env proteins also formed in these baculovirus-infected insect cells. Immunofluorescence staining of intact insect cells infected by the baculovirus expressing BIV Env with BIV-specific serum demonstrated that the recombinant Env glycoproteins were expressed on the cell surface. Antisera raised to recombinant Env glycoproteins immunoprecipitated native gp145, gp100, and gp45 in BIV-infected bovine cells similar to sera from animals naturally or experimentally infected with BIV.

Animals↗
PubMed1992

Effects of the herbicides hexazinone and triclopyr ester on aquatic insects.

Experiments were conducted to measure acute lethal response of aquatic insects to hexazinone (Velpar L) and triclopyr ester (Garlon 4) in flow-through laboratory bioassays, and to determine lethal and behavioral effects of these herbicides on insects in outdoor stream channels. No significant mortality (chi 2 P greater than 0.05) occurred in 13 test species exposed to hexazinone in laboratory flow-through bioassays (1-hr exposure, 48-hr observation) at the maximum test concentration of 80 mg/liter. The survival of insects exposed to 80 mg/liter hexazinone in outdoor stream channels was likewise unaffected. Significant drift (chi 2 P less than 0.001) of Isonychia sp. occurred during a hexazinone treatment of the stream channels, but only at the maximum concentration of 80 mg/liter, and survival of the displaced Isonychia sp. was not affected. In flow-through bioassays with triclopyr ester, 10 of 12 test species showed no significant mortality at concentrations greater than 80 mg/liter. Survival of Isogenoides sp. and Dolophilodes distinctus was significantly affected at less than 80 mg/liter. Lethal concentrations were estimated by probit analysis of concentration-response data (1-hr exposure, 48-hr observation) for Simulium sp. (LC50 = 303 mg/liter), Isogenoides sp. (LC50 = 61.7 mg/liter), and D. distinctus (LC50 = 0.6 mg/liter). Triclopyr ester applications to the stream channels resulted in significant drift and mortality of D. distinctus at 3.2 mg/liter (no effects at 0.32 mg/liter), Isogenoides sp. at 32 mg/liter, and Hydropsyche sp. and Epeorus vitrea at 320 mg/liter. The risk to aquatic insects of these herbicides used in forest vegetation management is discussed.

Animals↗
PubMed1995

Visual ecology and voltage-gated ion channels in insect photoreceptors.

That particular membrane conductances are selected for expression to enable the efficient coding of biologically relevant signals is illustrated by recent work on insect photoreceptors. These studies exploit the richness of insect vision and the accessibility of insect photoreceptors to cellular analysis in both intact animal and isolated cell preparations. The distribution of voltage-gated conductances among photoreceptors of different species correlates with visual ecology. Delayed-rectifier K+ channels are found in the rapidly responding photoreceptors of fast-flying flies. The conductance's activation range and dynamics match light-induced signals, and enable a rapid response by reducing the membrane time constant. Slow-moving flies have slowly responding photoreceptors that lack the delayed rectifier, but express an inactivating K+ conductance that is metabolically less demanding. Complementing these findings, locust photoreceptor membranes are modulated diurnally. The delayed rectifier is exhibited during the day and the inactivating K+ current is exhibited at night. Insect photoreceptors also demonstrate the amplification of signals by voltage-gated Na+ channels. In drone-bee photoreceptors, voltage-gated Na+ channels combine with K+ channels to enhance the small transient signals produced by the image of a queen bee passing over the retina. This subthreshold amplifier operates most effectively over the range of light intensities at which drones pursue queens.

Animals↗
PubMed1986

Effect of host insect sterols on the development and sterol composition of Steinernema feltiae.

Steinernema feltiae (= Neoaplectana carpocapsae), 'All' strain, was propagated in larvae of the corn earworm, Heliothis zea, which contained various sterols, in order to determine how the sterol composition of the host affects the growth, development and sterol composition of this insect-parasitic nematode. S. feltiae completed its life cycle normally in insects containing primarily cholesterol, cholestanol or 7-dehydrocholesterol, although the sterol composition of the dauer stage was affected by the sterol composition of the host. When the nematode was reared in insects containing primarily cholesterol, 55% of the sterol in the dauers was cholesterol and the other 46% was lathosterol. In contrast, cholestanol (70%) and lathostetrol (31%) were the sterols present in nematodes reared in H. zea containing primarily cholestanol. Cholestanol (43%), lathosterol (34%), campestanol and/or another 24-methylsterol (23%) and cholesterol (1%) were the sterols present in nematodes reared in H. zea containing campestanol and cholestanol as its major sterols. Lathosterol was the major sterol present in nematodes reared in H. zea containing principally 7-dehydrochlesterol. Therefore, in each case, S. feltiae metabolized some host sterol to lathosterol but the relative percentage of lathosterol in the nematode increased as it was exposed to delta 0-, delta 5- and delta 5,7-sterols, respectively. The ability of S. feltiae to utilize different host sterols may, in part, explain its success in parasitizing a wide variety of insects.

Animals↗
PubMed1992

Insect defensins: inducible antibacterial peptides.

In response to bacterial challenge or trauma, insects produce a battery of bactericidal or bacteriostatic molecules with a broad spectrum of activity against Gram-positive and/or Gram-negative bacteria; most are small-sized cationic peptides. This review focuses on insect defensins, a large group of inducible antibacterial peptides that are present both in ancient and recent insect orders. This immune response of insects shares many of the characteristics of the mammalian acute phase response.

Amino Acid Sequence↗
PubMed1996

Purification and in vitro-phospholabeling of secretory envelope proteins E1 and E2 of hepatitis C virus expressed in insect cells.

The putative envelope glycoproteins of hepatitis C virus (HCV), E1 and E2, were expressed as recombinant, secretory proteins in Sf9 insect cells through infection with recombinant baculoviruses. The influenza virus hemagglutinin signal sequence (HASS) was inserted upstream of the HCV-cDNAs in order to effect secretion. Furthermore, a hexa-histidine tag for purification on a Ni(2+)-nitrilotriacetic acid (Ni(2+)-NTA) column and a protein kinase A (PKA) recognition sequence for in vitro-phospholabeling were fused upstream of the HCV-cDNA. E1- and E2 proteins lacking their carboxy-terminal, hydrophobic sequence were produced by baculovirus-infected insect cells in bioreactors of 23 1. The medium was concentrated and proteins were purified under native conditions on Ni(2+)-NTA columns. Purified proteins could be phospholabeled in vitro using the catalytic subunit of protein kinase. A isolated from bovine heart and gamma-[32P]ATP. Labeled E1 and E2 proteins expressed in insect cells could be immunoprecipitated with sera from HCV-infected patients. Co-expression of these E1 and E2 proteins led to the formation of E1-E2 complexes within the insect cell and to secretion of these complexes into the medium.

Adenosine Triphosphate↗
PubMed1988

Immune mechanisms in insects.

Understanding immune evasion by parasites in their insect vectors requires some understanding of the insect immune system. Until fairly recently, technical difficulties in handling cells and plasma hampered laboratory investigations into insect immunology, but modern techniques combined with a sound knowledge of insect physiology are now permitting rapid advances. Rather than discussing the many controversies, this review aims to point out current areas of research into cellular and 'humoral' mechanisms that might be followed up by parasitologists.

Journal Article↗
PubMed1994

The inducible antibacterial peptides of insects.

Insects respond to bacterial challenge by the rapid and transient synthesis of a large number of potent antibacterial peptides that are active against many different bacteria. Two families of inducible antibacterial peptides are well characterized: the cecropins and the insect defensins. A rapidly increasing number of proline- and glycine-rich peptides are reported from various insect species together with cecropins and insect defensins. In this review, Stéphane Cociancich, Philippe Bulet, Charles Hetru and Jules A. Hoffmann give an update of our current information on the induced antibacterial peptides.

Journal Article↗
PubMed1985

Neuropeptide immunocytochemistry in protostomian invertebrates, with special reference to insects and molluscs.

In some molluscs (Aplysia and Fusitriton) and insects (silkworm and cricket), occurrence and distribution of neuropeptides in the nervous system and gut were studied with following results: in these invertebrates and also in planaria, PP-like immunoreactivity is extensively distributed in neurons and (in insects) in gut endocrine paraneurons. These cells are negative for NPY, the mammalian neuropeptide related to PP in molecular structure. PHI-like immunoreactivity is widely distributed in the neurons of those invertebrates; it occurs also in gut endocrine paraneurons in insects. The PHI-immunopositive cells are immunonegative for VIP and the coexistence of both peptides due to the common precursor in mammals cannot be recognized in these invertebrates. Immunoreactivity for urotensin I, the neuropeptide derived from teleostean urophysial neurons, is widely distributed in the neurons of the invertebrates. In insects (cricket) it occurs in gut endocrine cells.

Animals↗
PubMed1987

Heavy metal levels in two biennial pine insects with sap-sucking and gall-forming life-styles.

The concentrations of cadmium, copper, nickel and lead were studied in two biennial pine insects in relation to the deposition of heavy metals in the environment around the industrialised town of Harjavalta in southwestern Finland. Sap-sucking pine bark bugs, Aradus cinnamomeus (Heteroptera, Aradidae), and gall-forming pine resin gall moths, Petrova resinella (Lepidoptera, Tortricidae) were collected on sample plots located at logarithmic distances along 9 km-long transects from the distinctive emission source. The responses of these insects representing different life-style were compared. Heavy metal concentrations in A. cinnamomeus were highest (Cd 17 microg g(-1), Cu 1900 microg g(-1), Ni 220 microg g(-1), Pb 32 microg g(-1)) in the vicinity of the factor complex, and lowest in the outermost zones. This trend followed a linear regression model. The pattern was less clear in P. resinella, the concentrations being only one-tenth of those recorded in A. cinnamomeus. Correlations between metal levels in A. cinnamomeus and previously examined Sphagnum moss bags proved to be highly significant in every case. The differences in the heavy metal concentrations of these two insect species, which occupy the same trophic position, would appear to be due to the differences in their feeding characteristics. Heavy metals accumulate in the posterior bulb of the midgut in the discontinuous alimentary system of A. cinnamomeus, while P. resinella is likely to secrete most of the metals into the walls of the galls. The almost total absence of these two insect species near the factory complex seems to be associated with the high concentrations of metals.

Journal Article↗
PubMed1991

An alternative explanation for anomalies in "soluble lipofuscin" fluorescence data from insects, crustaceans, and other aquatic species.

Published attempts to extract lipofuscin from crustaceans and fish to assess age for fisheries research purposes have used essentially the same extraction methodology applied to insects, but have neither shown a conclusive age-dependence of spectrally similar fluorescence nor proved its association with lipofuscin. The reported lipofuscin solvent extraction method for fleshflies, Sarcophaga bullata, was manipulated by varying wash volume. This revealed that almost all age-dependent blue fluorescent material persisting in lipid fractions was actually pteridine-like. This finding was consistent with some previous independent results for Musca domestica. Examination of reported lipofuscin extraction protocols for other insects suggested that this problem was probably widespread. The pteridines are known to occur in unusually high amounts in insects, accumulating with age specifically in some members of this group by storage excretion, probably as a terrestrial water conservation strategy. In addition, there is growing evidence in the gerontological literature for other groups that solvent extracted blue fluorescence is not a true measure of lipofuscin content in tissues. These findings provide considerable insight into anomalies in putative lipofuscin fluorescence data between the insects and various aquatic species and suggest that there may be little basis for expectations of age-dependent fluorescence from aquatic species when the same gross extraction and crude purification methods are used.

Animals↗
PubMed2006

Comparison of different signal peptides for protein secretion in nonlytic insect cell system.

Protein expression and secretion in insect cells have been widely studied in the baculovirus-infected insect cell system. In directly transfected insect cells only intracellular expression and purification of recombinant proteins have been studied in detail. To examine multiple recombinant protein variants, easy and fast expression and a purification screening system are required. The aim of this study was to establish an effective and rapid secretion system for human azurocidin using directly transfected insect cells. We also constructed and tested expression vectors possessing heterologous signal peptides derived from human azurocidin, yellow lupin diphosphonucleotide phosphatase/phosphodiesterase (PPD1), and papaya papain IV to secrete yellow lupin and red kidney bean purple acid phosphatases, PPD1, and papain IV. Our results demonstrate that the secretion vectors used here can direct recombinant proteins to the culture medium very effectively, allowing their simple purification on a small/medium scale. Based on secretion and activity analyses it seems that the azurocidin signal peptide is one of the most potent secretion signals.

Acid Phosphatase↗
PubMed2006

Absorption of toxic beta-glucosides produced by plants and their effect on tissue trehalases from insects.

Trehalases present in body wall, Malpighian tubules, fat body, midgut and haemolymph from Tenebrio molitor (Coleoptera), Musca domestica (Diptera), Spodoptera frugiperda and Diatraea saccharalis (Lepidoptera) were assayed in the presence and absence of toxic beta-glucosides produced by plants or their aglycones. The glucosides used were phlorizin, amygdalin, prunasin and the aglycone mandelonitrile. In addition, T. molitor and S. frugiperda trehalases were assayed with and without esculin. More than 60% of total trehalase activity was found in the midgut of these insects. As a rule, trehalases present in each insect were inhibited by at least two of the glucosides. Prunasin was the best inhibitor in tissues with highest trehalase activity. S. frugiperda beta-glucosidases were not able to hydrolyze esculin. Nevertheless, their larval midguts absorb the intact glucoside that is recovered from the fat body, Malpighian tubules and mainly from haemolymph. Mature larvae fed on a diet containing 3 mM (0.1%) esculin have 0.2 mM esculin in their haemolymph, and weigh 60% of control larvae. In vitro, haemolymph trehalase activity is abolished by 0.5 mM esculin. This inhibition may play a role in the decrease of body weight and in animal survival. S. frugiperda larvae reared in 0.1% amygdalin-containing diet present higher trehalase activity in tissues than the larvae reared in 0.1% esculin-containing diet. Higher trehalase activity should be the reason why the S. frugiperda development is not impaired by 1% dietary amygdalin, in contrast to what is observed when insects are reared in 0.1% esculin. The data suggest that many plant beta-glucosides are toxic because they inhibit trehalase, a key enzyme controlling glucose availability in insects.

Animals↗