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PubMed1997

Egg shells of mallophagans and anoplurans (Insecta: Phthiraptera): morphogenesis of specialized regions and the relation to F-actin cytoskeleton of follicular cells.

The egg shells of investigated phthirapterans consist of three basic elements: an anterior operculum, a main egg shell and a posterior hydropyle. In some species these elements show further regional specializations: bristles and projections that facilitate attachment to feathers of the host, micropyles and aeropylar openings. All of the egg shell specializations are formed by distinct subpopulations of follicular cells. Staining with rhodamine-conjugated phalloidin has revealed that these subpopulations significantly differ in the distribution of microfilaments (F-actin). In this respect four morphological categories of the follicular cells have been distinguished: (1) cells devoid of processes and microvilli, with basal arrays of microfilaments, responsible for the secretion of a flat chorion; (2) cells devoid of processes and microvilli, separated by intercellular spaces, with basal arrays of microfilaments, responsible for the secretion of attachment structures; (3) cells equipped with actin-containing processes, responsible for the formation of micropyles or aeropyles, and (4) cells equipped with bundles of microvilli, responsible for the formation of hydropyles.

Actins↗

PubMed1998

Immunohistochemical localization of ecdysteroid receptor and ultraspiracle in the epithelial cell line from Chironomus tentans (Insecta, Diptera).

Ecdysteroid receptor (EcR) and its heterodimerization partner, ultraspiracle (USP), were demonstrated in the epithelial cell line from Chironomus tentans by immunohistochemistry. In untreated cells both proteins are present in nuclei as well as in granular compartments of the cytosol. At 1 day after addition of 1-microM 20-OH-ecdysone (20E) total immunofluorescence had increased in the nuclei, whereas the cytoplasmic staining had disappeared. At the 2nd and 3rd days all cells within a vesicle appear identical according to morphological criteria, but the EcR and USP immunoreactivity becomes restricted into patches of neighbouring cells. The hormonally induced changes in the pattern of localization of functional ecdysteroid receptor, the heterodimer of EcR and USP, are discussed in relation to similar effects of 20E on acetylcholinesterase and muscarinic acetylcholine receptor distribution in this cell line.

Animal Structures↗

PubMed2003

Bioaccumulation and toxicity of sediment associated herbicides (ioxynil, pendimethalin, and bentazone) in Lumbriculus variegatus (Oligochaeta) and Chironomus riparius (Insecta).

The benthic macroinvertebrates Lumbriculus variegatus and Chironomus riparius were used in toxicity and bioaccumulation tests to determine the toxic concentrations and accumulation potential of sediment associated herbicides. The tested chemicals were ioxynil, bentazone, and pendimethalin. The bioaccumulation tests with L. variegatus were performed in four different sediments, each having different characteristics. Water-only LC(50) tests were performed with both L. variegatus and C. riparius. A sublethal effect of model compounds in sediments was assessed by a C. riparius larvae growth-inhibition test. Of the model compounds, ioxynil appeared to be the most toxic, with LC(50) values 1.79 and 2.79 mgL(-1) for L. variegatus and C. riparius, respectively. The LC(50) water concentrations for bentazone were 79.11 and 62.31 mgL(-1) for L. variegatus and C. riparius, respectively. Similarly, ioxynil revealed the highest bioaccumulation potential in bioaccumulation tests. The most important characters affecting chemical fate in the sediment seemed to be the organic matter content and the particle size fraction. The sediments with low organic material and coarse particle size consistently showed high bioaccumulation potential and vice versa. In C. riparius growth tests bentazone had a statistically significant effect on larval growth at sediment concentrations of 1160 and 4650 mgkg(-1) (P<0.05). It is noteworthy that standard deviations tend to be greater at high chemical concentrations, which addresses the fact that part of the individuals started to suffer. Ioxynil had an effect on the larval growth in other test sediment at the highest concentration (15.46 mgkg(-1)dw), in which head capsule length correlated with larval weight, decreasing toward higher exposure concentrations. The current results show the importance of sediment organic matter as a binding site of xenobiotics.

Aniline Compounds↗

PubMed2002

Expression of ecdysteroid receptor and ultraspiracle from Chironomus tentans (Insecta, Diptera) in E. coli and purification in a functional state.

Full length clones of ecdysteroid receptor (EcR) and Ultraspiracle (USP) from Chironomus tentans were expressed as GST fusion proteins in E. coli and purified by affinity chromatography. The absence of detergents during the purification procedure is essential for retaining receptor function, especially ligand binding. Presence of USP is mandatory for ligand binding to EcR, but no other cofactors or posttranslational modifications seem to be important, since Scatchard plots revealed the same characteristics (two high affinity binding sites for Ponasterone A with K(D1)=0.24+/-0.1nM and K(D2)=3.9+/-1.3.nM) as found in 0.4 M NaCl extracts of Chironomus cells. Gel mobility shift assays showed binding of the heterodimer to PAL and DR5 even after removal of the GST-tag, whereas EcR binding to PAL1 is GST-dependent. USP binds preferentially to DR5. Addition of unprogrammed reticulocyte lysate improves ligand binding only slightly. Removal of GST has no effect on (3)H-ponasterone A binding, but alters DNA binding characteristics. Calculation of specific binding (5.3+3.0 nmol/mg GST EcR) revealed that 47+/-26% of purified receptor protein was able to bind ligand. The addition of purified EcR to cell extracts of hormone resistant subclones of the epithelial cell line from C. tentans, which have lost their ability to bind ligand, restores specific binding of (3)H-ponasterone A.

Acrylic Resins↗

PubMed1998

Ecdysteroid receptor and ultraspiracle from Chironomus tentans (Insecta) are phosphoproteins and are regulated differently by molting hormone.

Three different isotypes of the ecdysteroid receptor (cEcR) (66, 68 and 70 kDa) and several molecular variants of the dimerization partner "ultraspiracle" (cUSP) (58-77 kDa) can be separated electrophoretically in homogenates of the epithelial cell line from Chironomus tentans. After phosphatase treatment the bands with the lowest electrophoretic mobility disappear in both cases. Phosphorylation occurs exclusively at ser/thr in EcR and USP. Binding studies with 3H-ponasterone A using 0.4 M NaCl extracts revealed two classes of high-affinity binding (KD1 = 0.47 and KD2 = 7.2 nM) competable either with 20-OH-ecdysone or muristerone A. At least KD2 and Bmax2 are unchanged after dephosphorylation. In hormonally naive cells a considerable part of EcR and USP is already present in nuclei. The phosphorylation pattern of both transcription factors is the same in cytosol and nuclear fractions. Incubation with 20-OH-ecdysone (1 microM, up to 4 days) does not alter the extent and mode of phosphorylation of EcR, although EcR concentration increases. In contrast USP concentration remains constant but phosphorylation is enhanced.

Animal Structures↗

PubMed1998

Cooperation of dopachrome conversion factor with phenoloxidase in the eumelanin pathway in haemolymph of Locusta migratoria (Insecta).

Dopachrome Conversion Factor (DCF) was found in the plasma of the locust Locusta migratoria. It has an apparent molecular mass of 85,000. Its K(m) was 0.2 mM at 22 degrees C and pH 7 with L-dopachrome as substrate. It had a high substrate specificity for L-dopachrome, methyl-L-dopachrome and L-dopachrome methyl ester but no activity on the corresponding D-isomers or on dopaminechrome. DCF was devoid of any phenoloxidase activity. Under action of DCF, L-dopachrome was converted into dihydroxyindole, which showed that a decarboxylation occured in the course of reaction. Locust DCF was inhibited by indole-3-propionic acid but not by indole-3- or indole-2-carboxylic acid. It was also inhibited by L-tryptophan in a competitive manner. Inhibition and substrate specificity suggest that a carboxyl group, either free or as a methyl ester, is necessary but not sufficient for enzyme recognition. When purified prophenoloxidase was activated and then added to dihydroxyindole either prepared by chemical synthesis or obtained by action of purified DCF on dopachrome, black pigments with a maximum absorption at 540 nm were generated. Therefore in the eumelanin pathway of locust plasma, phenoloxidase can catalyze the reaction that converts the product generated by DCF.

Animals↗

PubMed2001

Ergastoplasmic paracrystalline inclusion bodies in the adipose gonadal envelope and fat body of the glow worm, Lampyris noctiluca (Insecta, Coleoptera).

The gonads of glow worm larvae are enveloped by adipose tissue which represents a specialized fat body. The adipose gonadal envelope, and also to a lesser extent the fat body cells, contain tubular paracrystalline inclusion bodies (PIBs). Cells of other tissues are devoid of such inclusions. The PIBs form in the cisternae of rough ER. In young larvae PIB formation is sparse, but at advanced larval stages PIBs often occur as bundles in stacks of ergastoplasm. Typically, a PIB within a cisterna consists of four to seven parallel tubules. The outer diameter of a tubule is ca 28.8 nm and the width of the tubule lumen ca 12.2 nm. The "wall" of a tubule contains globular protein subunits of ca 8.3 nm diameter; the subunits are arranged helically. Since the adipose gonadal envelope progresses through a cytological differentiation process during differentiation and maturation of the gonads, the increased number of PIBs may indicate enhanced metabolic activity of the tissue related to nutrition of the growing gonads.

Adipose Tissue↗

PubMed2003

Dramatically elevated rate of mitochondrial substitution in lice (Insecta: Phthiraptera).

Few estimates of relative substitution rates, and the underlying mutation rates, exist between mitochondrial and nuclear genes in insects. Previous estimates for insects indicate a 2-9 times faster substitution rate in mitochondrial genes relative to nuclear genes. Here we use novel methods for estimating relative rates of substitution, which incorporate multiple substitutions, and apply these methods to a group of insects (lice, Order: Phthiraptera). First, we use a modification of copath analysis (branch length regression) to construct independent comparisons of rates, consisting of each branch in a phylogenetic tree. The branch length comparisons use maximum likelihood models to correct for multiple substitution. In addition, we estimate codon-specific rates under maximum likelihood for the different genes and compare these values. Estimates of the relative synonymous substitution rates between a mitochondrial (COI) and nuclear (EF-1alpha) gene in lice indicate a relative rate of several 100 to 1. This rapid relative mitochondrial rate (>100 times) is at least an order of magnitude faster than previous estimates for any group of organisms. Comparisons using the same methods for another group of insects (aphids) reveals that this extreme relative rate estimate is not simply attributable to the methods we used, because estimates from aphids are substantially lower. Taxon sampling affects the relative rate estimate, with comparisons involving more closely related taxa resulting in a higher estimate. Relative rate estimates also increase with model complexity, indicating that methods accounting for more multiple substitution estimate higher relative rates.

Animals↗

PubMed2003

Phylogenetic position of Phthiraptera (Insecta: Paraneoptera) and elevated rate of evolution in mitochondrial 12S and 16S rDNA.

Phthiraptera (chewing and sucking lice) and Psocoptera (booklice and barklice) are closely related to each other and compose the monophyletic taxon Psocodea. However, there are two hypotheses regarding their phylogenetic relationship: (1) monophyletic Psocoptera is the sister group of Phthiraptera or (2) Psocoptera is paraphyletic, and Liposcelididae of Psocoptera is the sister group of Phthiraptera. Each hypothesis is supported morphologically and/or embryologically, and this problem has not yet been resolved. In the present study, the phylogenetic position of Phthiraptera was examined using mitochondrial 12S and 16S rDNA sequences, with three methods of phylogenetic analysis. Results of all analyses strongly supported the close relationship between Phthiraptera and Liposcelididae. Results of the present analyses also provided some insight into the elevated rate of evolution in mitochondrial DNA (mtDNA) in Phthiraptera. An elevated substitution rate of mtDNA appears to originate in the common ancestor of Phthiraptera and Liposcelididae, and directly corresponds to an increased G+C content. Therefore, the elevated substitution rate of mtDNA in Phthiraptera and Liposcelididae appears to be directional. A high diversity of 12S rDNA secondary structure was also observed in wide range of Phthiraptera and Liposcelididae, but these structures seem to have evolved independently in different clades.

Animals↗

PubMed2004

Evolution and phylogenetic utility of CAD (rudimentary) among Mesozoic-aged Eremoneuran Diptera (Insecta).

We sequenced nearly the entire carbomoylphosphate synthase (CPS) domain of CAD, or rudimentary, (ca. 4 kb) from 29 species of flies representing all major clades within Eremoneura, or higher flies, and several orthorrhaphous brachyceran outgroups. We compared these sequences with orthologs from Anopheles gambiae and Drosophila melanogaster to assess structure, compositional bias, and phylogenetic utility. CAD is large (6.6+ kb), complex (comprised of three major and myriad minor functional domains) and relatively free of introns, extreme nucleotide bias (except third codon positions), and large hypervariable regions. The CPS domain possesses moderate levels of nonsynonymous divergence among taxa of intermediate evolutionary age and conveys considerable phylogenetic signal. Phylogenetic analysis of CPS sequences under varying methods and assumptions resulted in well-resolved, strongly supported trees concordant with many traditional ideas about higher dipteran phylogeny and with prior inferences from 28S rDNA. The most robustly supported major eremoneuran clades were Cyclorrhapha, Platypezoidea, Eumuscomorpha, Empidoidea, Atelestidae, Empidoidea exclusive of Atelestidae, Hybotidae s.l., Microphoridae+Dolichopodidae, and Empididae s. str. Because CAD is ubiquitous, apparently single copy (at least within holometabolous insects), readily obtained from several insect orders using primers described herein, and exhibits considerable phylogenetic utility, it should have wide applicability in insect molecular systematics.

Animals↗

PubMed2001

Arachidonic and eicosapentaenoic acids in tissues of the firefly, Photinus pyralis (Insecta: Coleoptera).

We report on the presence of high proportions of arachidonic acid (20:4n-6) and eicosapentaenoic acid (20:5n-3) in the tissue lipids of adult fireflies, Photinus pyralis. Arachidonic acid typically occurs in very small proportions in phospholipids (PLs) of terrestrial insects, ranging from no more than traces to less than 1% of PL fatty acids, while 20:5n-3 is often missing entirely from insect lipids. Contrarily, 20:4n-6 made up approximately 21% of the PL fatty acids prepared from whole males and females, and from heads and thoraces prepared from males. Proportions of 20:4n-6 associated with PLs varied among tissues, including approximately 8% for male gut epithelia, 13% for testes, and approximately 25% for light organs and body fat from males. Substantial proportions of 20:5n-3 were also associated with PLs prepared from male firefly tissues, including 5% for body fat and 8% for light organs. Because 20:4n-6 and 20:5n-3 are precursors for biosynthesis of prostaglandins and other eicosanoids, we considered the possibility that firefly tissues might produce eicosanoids at exceptionally high rates. Preliminary experiments indicated otherwise. Hence, fireflies are peculiar among terrestrial insects with respect to maintaining high proportions of PL 20:4n-6 and 20:5n-3.

Animals↗

PubMed2001

Cryoprotective role of polyols independent of the increase in supercooling capacity in diapausing adults of Pyrrhocoris apterus (Heteroptera: Insecta).

Diapausing cold-acclimated adults of the bug Pyrrhocoris apterus accumulate four 'winter' polyols, ribitol, sorbitol, mannitol and arabinitol, in total concentrations of up to 100 mM. The accumulation started only when the temperatures dropped below a threshold of +5 degrees C in laboratory acclimated insects. The supercooling capacity (SCP) was not affected by polyol accumulation and remained constant at approximately -17 degrees C. Cold hardiness, measured as survival time (Lt50) at -15 degrees C, increased from approximately 1 day to approximately 1 week in parallel with polyol accumulation. There was a tight correlation (r=0.98) between the concentration of 'winter' polyols in haemolymph and Lt50(-15). When a mixture of ribitol and sorbitol was injected into the haemolymph of the bugs acclimated to +5 degrees C, the concentration of polyols increased from 2.5 to 83.1 mM in haemolymph, or from 0.07 to 6.61 microg/mg of fresh weight in the whole body, the SCP remained unchanged and survival after exposure to -14 degrees C for 3 days increased approximately three-fold in comparison to untreated controls. Such results were interpreted as evidence for the cryoprotective role of accumulated polyols independent of the depression of SCP. Acclimation protocol using thermoperiod, mimicking daily temperature oscillations, resulted in moderately lower SCP, higher sum of polyols accumulated and significantly longer Lt50(-15) than at acclimation protocol with constant temperatures.

Adaptation, Physiological↗

PubMed1976

Production of larval tanypodinae (Insecta: Chironomidae) in the mud at Loch Leven, Kinross.

Life histories and production of the five common species of Tanypodinae (Pentaneura monilis, Procladius simplicistilus, P. crassinervis, P. choreus and Psilotanypus rufovittatus) in the mud at Loch Leven were measured from March 1971 to March 1972. The methods used to identify each species of larva are described. All were univoltine except Pentaneura monilis which had two generations per year, but the seasonal patterns of the life cycles were different. The average density of third and fourth instar Tanypodinae was 3100/m2 and densities of individual species exceeded 2500/m2 for short periods in Procladius crassinervis and P. choreus. Net annual dry weight production by this subfamily was 2-6 g/m2 (57 KJ/m2), with individual species ranging from 1-1 g/m2 (P. crassinervis) to 0-16 g/m2 (Pentaneura monilis). These results are discussed briefly and compared with production estimates for other zoobenthos.

Animals↗

PubMed2000

Purification and characterization of an esterase conferring resistance to fenitrothion in Oryzaephilus surinamensis (L.) (insecta, coleoptera, silvanidae).

Esterases from a fenitrothion-resistant strain (VOSF) of the saw-toothed grain beetle, Oryzaephilus surinamensis (L.), are presumed to play a role in conferring resistance to malathion, fenitrothion, and chlorpyrifos-methyl. Colorimetric assays showed a significant positive correlation between increased resistance to fenitrothion in strains of O. surinamensis examined and elevated esterase hydrolytic activity to substrates of p-nitrophenyl acetate, alpha-naphthyl acetate, and beta-naphthyl acetate. Esterase zymograms showed different banding patterns between VOSF and an insecticide-susceptible strain, VOS48. A major esterase in the VOSF strain, not detected in VOS48, was purified and characterized by chromatographic and electrophoretic techniques. On the basis of SDS-polyacrylamide gel eletrophoresis, the molecular mass of the purified esterase from VOSF was 130 kDa and consisted of two 65 kDa subunits. Additional properties of this enzyme are discussed.

Animals↗

PubMed1999

Two new species of larval mites (Acari: Trombidioidea: Microtrombidiidae and Johnstonianidae) parasitising Culicoides impunctatus, the highland midge (Insecta: Ceratopogonidae), in Scotland.

The larval stage of the trombidioid mites Parafeiderium stuarti n. sp. (family Microtrombidiidae Thor, 1935) and Centrotrombidium blackwellae n. sp. (family Johnstonianidae Newell, 1957) are described. Both species were found parasitising Culicoides impunctatus Goetghebuer, the highland midge (Diptera: Ceratopogonidae), in Scotland. A revised diagnosis for Parafeiderium Vercammen-Grandjean & Cochrane, 1974 and a key to the world species of larval Centrotrombidium Kramer, 1896 are given.

Animals↗

PubMed2002

Principles of the highly ordered arrangement of metaphase I bivalents in spermatocytes of Agrodiaetus (Insecta, Lepidoptera).

We have investigated the nature of highly ordered bivalent arrangement in lepidopteran spermatocytes by analysing and comparing the patterns of bivalent distribution in intact metaphase I plates of 24 closely related species of the genus Agrodiaetus (Lycaenidae). The studied species greatly differed in haploid chromosome numbers (from n = 13 to n = 90) and in the structure of their karyotypes. We found that the larger the bivalent, the closer to the centre of the metaphase plate it was situated. In species with a high chromosome number and asymmetrical karyotype structure, the largest bivalent was located in the centre of the circular metaphase plate. Bivalents of equal size were approximately equidistant from the centre of the metaphase plate and formed concentric circles around the largest bivalent. These principles are diametrically different from those known in the majority of other animals and plants, in which the smallest elements of the chromosome set are situated in the centre of metaphase plate. The only exception from the above principles was observed in spermatocytes of A. surakovi which were heterozygous for reciprocal translocation involving two or three chromosome pairs. In addition to one large bivalent, the heterozygous cells had a multivalent, the size of which was comparable to or even exceeded that of the largest bivalentin the karyotype. In spite of thelarge size, the multivalent was always situated at the periphery of metaphase plate. This indicated that the chromosome size itself is not the only factor determining the bivalent position. We also found that the structure of the metaphase plate is fundamentally different in mitotic and meiotic cells of Agrodiaetus. In spermatogonial metaphase, chromosomes were tightly brought together, forming a dense compact disk, whereas during metaphase I of spermatocytes, all bivalents were clearly separated from each other, and the distance between adjacent bivalents varied from 0.4 to 1.5 microm. Based on the above findings, we proposed a model of bivalent distribution in the Lepidoptera. According to the model, during congregation in the prometaphase stage there is a centripetal movement of bivalents made by a force directed to the centre of the metaphase plate transverse to the spindle. This force is proportional to the kinetochore size of a particular bivalent. The Lepidoptera have a special near-holokinetic type of chromosome organisation. Therefore, large bivalents having large kinetochores are situated in the central part of metaphase plate. Another possible factor affecting the bivalent position is the interaction of bivalents with the cisternae of the membrane system compartmentalising the intraspindle space.

Animals↗

PubMed1997

Sex chromosome differentiation in some species of Lepidoptera (Insecta).

Sex chromosome morphology of eight Lepidoptera species was studied, exploiting predominantly the pachytene stage when chromosomes display a remarkable chromomere pattern. Six species had a WZ/ ZZ sex chromosome system, one species a W1W2Z/ ZZ system and one species was of the Z/ZZ type. Much like XY chromosomes in groups with male heterogamety, the lepidopteran sex chromosomes showed various degrees of structural differentiation. Differences between Z and W chromomere patterns ranged from undetectable to obviously non-homologous. A common property of the W chromosomes (the W1 in the W1W2Z/ZZ system) was the possession of a block of heterochromatin. The heterochromatin block comprised a small or a large segment of the W or even the entire W, depending on the species. Segments with apparent structural homology are evolutionarily young parts of the sex chromosomes-recently fused autosomes that have not had sufficient time for differentiation. The 'primitive' lepidopteran species Micropterix calthella had a Z/ZZ sex chromosome system. This supports the hypothesis that the lepidopteran W chromosome came into being at the base of the 'advanced' Lepidoptera; it was presumably an autosome whose homologue fused to the original Z chromosome.

Animals↗

PubMed2005

The head and body lice of humans are genetically distinct (Insecta: Phthiraptera, Pediculidae): evidence from double infestations.

Little is known about the population genetics of the louse infestations of humans. We used microsatellite DNA to study 11 double infestations, that is, hosts infested with head lice and body lice simultaneously. We tested for population structure on a host, and for population structure among seven hosts that shared sleeping quarters. We also sought evidence of migration among louse populations. Our results showed that: (i) the head and body lice on these individual hosts were two genetically distinct populations; (ii) each host had their own populations of head and body lice that were genetically distinct to those on other hosts; and (iii) lice had migrated from head to head, and from body to body, but not between heads and bodies. Our results indicate that head and body lice are separate species.

Animals↗