Cryopreservation of the late stage embryos of Spodoptera exigua (Lepidoptera: Noctuidae)

Genetic devolution, genetic drift and contamination are all threats to maintain germplasm stability during mass rearing of many insects. Cryopreservation of beet armworm (Spodoptera exigua) embryos was studied to provide information to improve mass rearing. A series of experiments was conducted on late-stage embryos (45-48 h at 27 degree C) of the beet armyworm, which included evaluation of cryoprotectants (CPAs), their toxicity and glass-forming tendency and optimization of experimental procedures. The results showed that ethylene glycol (EG) was the best CPA with comparatively low toxicity compared to the other six CPAs tested (methanol, 1,3-propanediol, glycerol, 2-amino-1-ethanol, 3-amino-1-propanol 3-methoxy-1 and 2-propanediol). The highest hatching rate of 8.8 degree was attained after freezing with a 3-step loading procedure and a 1-step unloading procedure, but the hatched larvae from frozen-thawed embryos did not actively feed and could not develop to a later stage. This was attributed to injuries from freezing in late stage embryos of S. exigua which had formed midguts.     

Ultrastructure of the Alabama argillacea (Hübner) (Lepidoptera: Noctuidae) midgut

The insect midgut has ultimately been the focus of researches tempting to control insect pests because alterations in the insect gut may affect not only its development, but also physiological events such as nutrient absorption and transformation. The objective of the present work was to describe morphologically, histochemically, and ultrastructurally the larva midgut of Alabama argillacea (Hübner) (Lepidoptera: Noctuidae), a cotton key pest in Brazil. Light and electronic transmission microscopy was used to obtain images from midgut sections of late fourth-instar larvae of A. argillacea. In general, the morphology, histochemistry, and ultrastructure characteristics of A. argillacea midgut follow that described in the literature for other lepidopteran species. However, the results showed a mitochondrial polymorphism and branched microvilli, which suggest an ultrastrucutural and physiological modification possibly associated with a high absorption and secretion activity by the columnar cells of this species. This intense activity may favor a faster response related to the action of ingested microbial agents and/or toxins, and can explain the high susceptibility of A. argillacea to the agents of control such as the toxin of Bacillus thuringiensis.     

Cold tolerance of field-collected and laboratory reared larvae of Sesamia nonagrioides (Lepidoptera: Noctuidae)

Sesamia nonagrioides Lefébvre (Lepidoptera: Noctuidae) is considered one of the most destructive pests of corn in the Mediterranean region. The purpose of the present study was to investigate some aspects of the cold tolerance of non-diapausing and diapausing laboratory reared larvae of S. nonagrioides, as well as of field-collected larvae, taking into consideration various parameters, such as supercooling ability, mean lethal temperature and accumulation of cryoprotectant substances, in relation to diapause. Our results provide evidence that S. nonagrioides has limited cold tolerance as it displays a low ability of supercooling. This is strongly supported by the fact that mortality of the individuals occurred after extended exposure to subzero temperatures, equivalent or slightly lower to their mean supercooling point. However, lethal temperatures of diapausing larvae were significantly lower in relation to that of non-diapausing larvae, indicating the existence of a direct link between diapause and cold tolerance. Regarding the role of cryoprotectant substances, accumulation of glycerol seems to be closely related to diapause, in contrast to accumulation of trehalose, which is more related to exposure to low temperatures slightly higher than 0 degree C. Finally, non-diapausing larvae of different instars displayed a similar ability of supercooling and tolerance to low temperatures as well as accumulation of cryoprotectant substances. The ecological significance of our findings on cold tolerance of this species is being discussed with particular reference to the microclimate observed in northern Greece.     

Changes in the haemocytes of Agrotis ipsilon larvae (Lepidoptera: Noctuidae) in relation to dimilin and Bacillus thuringiensis infections

Five types of haemocytes are observed in the fourth larval instar of the black cutworm, Agrotis ipsilon: prohaemocytes (PRs), plasmatocytes (PLs), granulocytes (GRs), spherule cells (SPs) and adipohaemocytes (ADs). Infection of A. ipsilon fourth larval instar with Bacillus thuringiensis and dimilin resulted in a reduction of the total haemocyte count. Changes in the differential haemocyte population during bacterial and dimilin infections have been assessed. The PRs % decreased significantly while SPs, PLs, and GRs % increased significantly after the application of the two insecticides at 12 and 24 h. Ultrastructural alternations and malformations have been observed in circulating haemocytes of A. ipsilon larvae treated with dimilin and B. thuringiensis.     

Microanatomy and ultrastructure of outer mantle epidermis of the cuttlefish,Sepia esculenta (Cephalopoda: Sepiidae)

This study describes the ultrastructural characteristics of external epidermis of mantle of Sepia esculenta using light and electron microscopy. The epidermis was thicker on the ventral surface than on the dorsal surface, with a higher secretory cell distribution on the ventral surface than on the dorsal surface. The epidermis was a single layer composed of epithelial cells, secretory cells, ciliated cells and neuroglial cells. Epithelial cells were columnar with well-developed microvilli on the free surface, and the microvilli were covered with glycocalyx. The epithelial cells were connected to the neighboring cells by tight junctions and membrane interdigitations of the apico-frontal surface. Well-developed microfilaments were arranged in a vertical direction in the cortical cytoplasm. The secretory cells were categorized into three types (A, B and C) in accordance with the light microscopical characteristics and ultrastructures of the secretory granules. The distribution of these cells was in the following order: Type A > Type B > Type C. SEM observation revealed that the secretory pore size of the Type A secretory cells was approximately 8.6 μm × 12.2 μm. Cytoplasm displayed a red color as the result of Masson's trichrome stain and H–E stain, and contained polygonal granules of approximately 1.2 μm² with a high electron density. The secretory pore size of the Type B secretory cells was approximately 10.1 μm × 12.1 μm. As the results of AB–PAS (pH 2.5) and AF–AB (pH 2.5) reactions, the cytoplasm displayed a red color. The cells contained membrane bounded secretory granules with very low electron density. The secretory pore of the Type C secretory cells was circular shape, and approximately 5.5 μm × 5.5 μm. Cytoplasm was found to be homogeneous under H–E stain and Masson's trichrome stain, and displayed a red color. As the result of AB–PAS (pH 2.5) reaction, the cytoplasm displayed a red color. The electron density of the secretory substance was the highest among the three types of secretory cells. The ciliated cells had a ciliary tuft on the free surface and were distributed throughout the mantle with the exception of the adhesive organs. Neuroglial cells were connected to the basal membrane, epithelial cells, secretory cells and nerve fibers through cytoplasmic process, and contained neurosecretory granules with high electron density within the cytoplasm.     

Circulating hemocytes from larvae of Melipona scutellaris (Hymenoptera,Apidae, Meliponini): Cell types and their role in phagocytosis

Infection in insects stimulates a complex defensive response. Recognition of pathogens may be accomplished by plasma or hemocyte proteins that bind specifically to bacterial or fungal polysaccharides. Several morphologically distinct hemocyte cell types cooperate in the immune response. Hemocytes attach to invading organisms and then isolate them by phagocytosis, by trapping them in hemocyte aggregates called nodules, or by forming an organized multicellular capsule around large parasites. In the current investigation the cellular in the hemolymph third instar larvae of M. scutellaris has been characterized by means of light microscopy analysis and phagocytosis assays were performed in vivo by injection of 0.5 μm fluorescence beads in order to identify the hemocyte types involved in phagocytosis. Four morphotypes of circulating hemocytes were found in 3rd instar larvae: prohemocytes, plasmatocytes, granulocytes and oenocytoids. The results presented plasmatocytes and granulocytes involved in phagocytic response of foreign particles in 3rd instar larvae of M. scutellaris.     

Fat body of Prorhinotermes simplex (Isoptera: Rhinotermitidae): ultrastructure, inter-caste differences and lipid composition

Ultrastructure of the fat body was studied in following castes and developmental stages of Prorhinotermes simplex: larvae of the first and the second instar, pseudergates, presoldiers, soldiers, nymphs, imagoes and mature ergatoid neotenic reproductives of both sexes. Fat body always consists of two principal cell types: adipocytes and urocytes. Adipocytes are characterized by a presence of large amounts of storage substances, namely lipid droplets, glycogen rosettes and proteins in the form of either biocrystals or vacuoles. Proportion of these components strongly varies during ontogeny. Adipocytes are equipped by a large central vacuole in which lipid droplets may resolve. Cytoplasm of urocytes contain glycogen rosettes and spherical or irregular concretions, other organelles are rare. Only adipocytes change their inner structure in the course of ontogeny: amount of glycogen decreases during the postembryonic development, it is the major kind of inclusion in the larvae but lacks in nymphs and imagoes; opposite trend is performed by lipids. The changes in protein content are less obvious but are explained and discussed. The total amount of triacylglycerols (TAGs) was found to be roughly 100microg in a pseudergate, 250microg in a nymph, and 30microg in a soldier. The most abundant fatty acids in TAGs are oleic (O), stearic (S), palmitic (P) and linoleic (L) acid. TAGs form a complex mixture with OOO, OPO, OLO and OOS being the most abundant isomers. Only negligible differences exist among the castes.     

Phagocytosis of latex beads and bacteria by hemocytes of the triatomine bug Rhodnius prolixus (Hemiptera: Reduvidae)

Insect circulating hemocytes are primarily responsible for the immune defense against parasites and pathogens. Here, we have analyzed phagocytosis of both biotic (bacteria) and abiotic (latex) particles by circulating hemocytes of 5th-instar nymphs of the triatomine bug Rhodnius prolixus. The following hemocyte types were identified: prohemocytes, plasmatocytes, granulocytes, oenocytoids and adipohemocytes. There was a considerable change in the relative percentage of plasmatocytes and prohemocytes in the hemolymph after challenge with both latex beads and bacteria. Granulocytes and oenocytoids also change their relative percentage in response to latex bead and Staphylococcus aureus, respectively. No significant change was observed in adipohemocytes at any time or treatment. Our data demonstrated that plasmatocytes were the only cell type involved in phagocytosis of foreign particles. As in mammal cells, phagocytosis by both zipper and trigger mechanisms were observed for the uptake of latex beads and bacteria. Neither melanization nor micro-aggregation was observed towards latex particles or Escherichia coli. On the other hand, R. prolixus produced a strong melanization reaction against S. aureus, thus showing that differences exist in the responses to E. coli and to S. aureus. Ultrastructural changes observed in plasmatocytes, adipohemocytes and oenocytoids suggest that these hemocyte types are directly involved in the immune defense of R. prolixus against foreign particles.     

Ultrastructure of the sensilla on larval antennae and mouthparts in the peach fruit moth, Carposina sasakii Matsumura (Lepidoptera: Carposinidae)

The ultrastructure of the sensilla on the larval antennae and mouthparts of the peach fruit moth, Carposina sasakii Matsumura, were investigated using scanning electron microscopy. On each three-segmented antenna, the basal scape is devoid of sensilla, the pedicel possesses two sensilla chaetica and three sensilla basiconica, and the terminal flagellum carries distally one sensillum styloconicum and three sensilla basiconica. The labrum bears six pairs of sensilla chaetica; the epipharynx carries two pairs of small epipharyngeal sensilla, three pairs of flattened sensilla chaetica, and a pair of broad sensillum digitiformium. Each dentate mandible carries two sensilla chaetica basally on its outer face. On the maxillae, the cardo and stipes each carry a sensillum chaeticum distally; each galea has three elongate flattened sensilla basiconica, two short sensilla chaetica, and two large sensilla styloconica. The distal segment of the maxillary palp possesses distally seven sensilla basiconica and one sensillum styloconicum, and laterally one sensillum digitiformium and one sensillum placodeum. Each labial palp bears distally a short cone-shaped sensillum chaeticum and an elongate slender sensillum styloconicum. The possible functions of these sensilla in the feeding process are briefly discussed.     

Effect of oil extraction assisted by ultrasound on the physicochemical properties and fatty acid profile of pumpkin seed oil (Cucurbita pepo)

The effects of amplitude and time of ultrasound-assisted extraction on the physicochemical properties and the fatty acid profile of pumpkin seed oil (Cucurbita pepo) were evaluated. Ultrasound time (5–30 min) and the response variables amplitude (25–100%), extraction yield, efficiency, oxidative stability in terms of the free fatty acids (FFA) of the plant design comprising two independent experiments variables, peroxide (PV), p-anisidine (AV), totox value (TV) and the fatty acid profile were evaluated. The results were analyzed by multiple linear regression. The time and amplitude showed significant differences (P < 0.05) for all variables. The highest yield of extraction was achieved at 5 min and amplitude of 62.5% (62%). However, the optimal ultrasound-assisted extraction conditions were as follows: ultrasound time of 26.34 min and amplitude of 89.02%. All extracts showed low FFA (2.75–4.93% oleic acid), PV (1.67–4.68 meq/kg), AV (1.94–3.69) and TV (6.25–12.55) values. The main fatty acids in all the extracts were oleic and linoleic acid. Therefore, ultrasound-assisted oil extraction had increased performance and reduced extraction time without affecting the oil quality.     

Comparative ultrastructure of spermatozoa of the redclaw Cherax quadricarinatus and the yabby Cherax destructor (Decapoda,Parastacidae)

Ultrastructure of spermatozoa of redclaw Cherax quadricarinatus and yabby Cherax destructor were described and compared. The acrosome complex and nucleus are located at the anterior and posterior region of the spermatozoon, respectively. The acrosome is a complex vesicle divided into two parts: the main body of the acrosome appears as a dense cup-shaped structure in longitudinal sagittal view, with the subacrosome zone occupying the central area of the vesicle. The acrosome is larger in C. quadricarinatus (width 2.37 ± 0.27 μm, length 1.31 ± 0.23 μm) than in C. destructor (width 1.80 ± 0.27 μm, length 1.01 ± 0.15 μm). There was no significant difference in L:W ratios of the studied species. The subacrosome zone in both species consists of two areas of different electron density. The nucleus is substantially decondensed and irregular in shape, with elaborate extended processes. The examined species exhibited a well-conserved structure of crayfish spermatozoon, similar to those of Cherax cainii and Cherax albidus. Small acrosome size, the absence of radial arms, and an extracellular capsule seem to be the morphological features that mostly distinguish Cherax from the Astacidae and Cambaridae.     

Antennal morphology and sensilla ultrastructure of the web-spinning sawfly Acantholyda posticalis Matsumura (Hymenoptera: Pamphiliidae)

Acantholyda posticalis (Hymenoptera: Pamphiliidae) is an important pine pest with a world-wide distribution. To clarify the olfactory receptive mechanism of A. posticalis, scanning electron microscopy and transmission electron microscopy were used to examine the morphology, ultrastructure, and distribution of antennal sensilla of adults from two sites in China. The antennae were filiform, and the flagella comprised 32–35 flagellomeres. Six sensillum types were found. Sensilla chaetica were straight setae with sharply pointed tips and without dendrites in the lumen. Sensilla trichodea were characterized by a parallel-grooved wall and one terminal pore and were innervated by four dendrites at the base. Sensilla basiconica I possessed longitudinally grooved surfaces and multiple terminal pores, with five dendrites in the lumen. Sensilla basiconica II not only had a distinct terminal pore but also had numerous tiny wall pores and many dendritic branches within the sensillum lymph. Sensilla coeloconica had deep longitudinal grooves, one terminal pore and six dendrites, while sensilla campaniformia were thick-walled with a terminal opening and sensory nerve bundles in the lumen. Sensilla chaetica and s. trichodea were most abundant and distributed over the entire antennae, while s. basiconica I and II, s. coeloconica, and s. campaniformia were restricted to the ventral flagellar surfaces. Although the shape and structure of antennae were similar in males and females, females had significantly longer antennae than males, and males had significantly more s. basiconica I than females. We compared the morphology and structure of these sensilla to other Hymenoptera and discussed their possible functions.     

Morphology and ultrastructure of the venom apparatus in the endoparasitic wasp Pteromalus puparum (Hymenoptera: Pteromalidae)

The venom apparatus of the endoparasitic wasp Pteromalus puparum (Hymenoptera: Pteromalidae) was studied with light and electron microscope and was subjected to the electrophoretic and immunohistochemical analyses. Typically its venom apparatus consists of an unbranched venom gland and a venom reservoir, which is associated with a Dufour gland. The venom gland is lined by a series of secretory units. Each secretory unit comprises a secretory cell and a duct cell. The secretory cell is associated with an end apparatus to collect its secretions into the gland lumen. Secretory cells in the venom gland are characterized by extensive rough endoplasmic reticulum and numerous electron-dense vesicles in the distal and middle parts. They also exhibit several secretory granules and vacuoles. The venom reservoir presents three distinct regions: an external layer, composed by numerous fine muscle fibers; an internal layer, represented by epithelial cell with large nucleus; and an intima portion, represented by thin and uniform organization. The morphological aspect of numerous well-developed organelles responsible for protein generation observed is in agreement with the electrophoretic and immunohistochemical results which reveal that the rich proteinaceous components are present in the venom gland and venom reservoir. The venom proteins are first mainly produced in the secretory unit of venom gland, then drained to the lumen through the end apparatus, and are finally collected and stored in the venom reservoir.     

Circulating hemocytes from larvae and adults of Carabus (Chaetocarabus) lefebvrei Dejean 1826 (Coleoptera, Carabidae): cell types and their role in phagocytosis after in vivo artificial non-self-challenge

Carabus lefebvrei Dejean 1826 is an helicophagous Italian endemic ground beetle that lives in central and south Apennines mountain forests, from lower altitudes to about 1500 m. In ground beetles, no morphofunctional data about immune system is available, even though they are well known both taxonomically and ecologically and they have been often used as indicators of the habitat quality due to their specificity to certain habitat types. In the current investigation the cellular population in the hemolymph of adult and third instar larvae of C. lefebvrei has been characterized by means of light and electron microscopy analysis and phagocytosis assays were performed in vivo by injection of 0.9 microm carboxylate-modified polystyrene latex beads in order to identify the hemocyte types involved in phagocytosis. Four morphotypes of circulating hemocytes were found both in larvae and in adults: prohemocytes, granulocytes, oenocytoids and plasmatocytes. After in vivo artificial non-self-challenge treatments, C. lefebvrei showed a non-specific immune response involving phagocytosis performed by plasmatocytes, both in adults and in larvae and by oenocitoids in larvae. In untreated animals, the hemocyte type presenting a firm phagocytic activity, the plasmatocytes, presented a percentage significantly higher in larvae than in adults, and after latex beads injections in larvae there was a tendency of significant difference in plasmatocyte percentage compared to controls injected with phosphate saline buffer. We think that these differences could be correlated with the peculiar morphology (less chitinization) and ecology of larval stages that are more sensitive to pathogens than adults.     

Structure and ultrastructure of the spermatozoa of Trypoxylon (Trypargilum) albitarse Fabricius 1804 (Hymenoptera: Apoidea: Crabronidae)

The ultrastructure of the spermatozoa of Trypoxylon (Trypargilum) albitarse is described here for the first time within this genus. Testes and seminal vesicles were dissected and processed for transmission electron microscopy. In the testicular follicles, the spermatids are arranged in a maximum number of 32 for each cyst. The spermatozoa are slender and they measure approximately 150 μm in length. The head is about 17 μm long and is formed by the acrosome and the nucleus. The flagellum consists in an axoneme, two mitochondrial derivatives, two accessory bodies and, at the nucleus–flagellum transition, a symmetric centriolar adjunct. The axoneme presents the typical 9 + 9 + 2 microtubule pattern. In the terminal region, the central microtubules and nine doublets finish first, followed by the accessory microtubules. Both mitochondrial derivatives begin together and are inserted in the base of the centriolar adjunct. Along the middle region, the larger derivative has almost twice the area of the smaller one and includes a discrete paracrystalline region. At the tip, the smaller derivative ends before the larger one and both before the axoneme. The characters derived from the ultrastructure of the spermatozoa of T. albitarse show synapomorphies shared with the Apoidea and present characters that are probably apomorphic for the Crabroninae subfamily.     

Effect of thermal treatment on the magnetic properties and structure of (Nd, Pr)-(Tb, Dy, Gd)-(Fe, Co, Al, Cu, Re)-type magnets

The effect of thermal treatment on the magnetic properties and structure of magnets made from an alloy of (wt %) 6.3 Nd, 6.84 Pr, 14.5 Dy, 2.12 Tb, 2.25 Gd, 14.0 co, 52.62 Fe, 0.08 Al, 0.03 Cu, 0.03 Re, 1.23 B is investigated. Following the optimum heat treatment (1175 K, 7.2 ks + cooling V = 0.02 K/s+ 675 K, 3.6 ks), baked magnets have the following magnetic properties at 300 K: B _r = 9.93 T, _j H _c = 1640 kA/m, H _k = 1360 kA/m, BH _max = 168 kJ/m³, α < |0.01|% K in the temperature range of 223–373 K. Nanoheterogenic areas 8–12 nm in size are found in grains of R ₂(Fe,M)₁₄B-type magnetosolid phase by means of atomic force microscopy.     

Morphology and ultrastructure of the hindgut fermentation chamber of a melolonthine beetle Holotrichia parallela (Coleoptera: Scarabaeidae) during larval development

The morphology and ultrastructure of the hindgut fermentation chamber of a melolonthine beetle, Holotrichia parallela, were examined using light and electron microscopy. The results showed that the anterior portion of the hindgut expanded into a characteristic lobe-like structure described as a fermentation chamber. Hematoxylin-eosin staining revealed that the wall of the fermentation chamber was composed of three main layers: the longitudinal muscle layer, the circular muscle layer, and the columnar epithelial cells. Scanning electron microscopy showed that the ultrastructure of the inner surface of the fermentation chamber was subjected to significant changes during larval development. Only some folds and a few cocci attached to the folds were found in the first-instar larvae. In the fermentation chambers of the second-instar larvae, a cuticular intima began to emerge, and the bacteria multiplied to form bacterial groups attached to the intima. A special lobe-like structure was formed in the third-instar larvae, constituted by bacteria and the bacteria-covered cuticular intima. Transmission electron microscopy revealed that the lobe-like structure held large numbers of rod-shaped bacteria. These data suggest that the hindgut fermentation chamber may have an important role in the symbiotic relationship between microbes and their insect hosts.     

Effect of pressure on the Kondo temperatures of Au(Fe) and Au(Mn)

The electrical resistivity of four Kondo systems, Au-(5 p.p.m. Fe), Au-(39 p.p.m. Fe), Au-(17 p.p.m. Mn) and Au-(50 p.p.m. Mn) has been measured in the temperature range 1.3–20 K at pressures up to 80 kbar. The Kondo temperature T_K is found to increase initially with pressure at the rate of 1.1%/kbar for Au(Fe) and 6%/kbar for Au(Mn). The volume dependence of the effective exchange constant J_eff is derived.     

溶剂效应对聚苯撑乙烯掺杂苝二酰亚胺太阳电池性能的影响

研究了不同溶剂对2-甲氧基-5-(2-乙基己氧基)-1，4-对苯撑乙烯(MEH-PPV): N,N′-二(1-乙基丙基)-3，4，9，10-苝四羧酸二亚酰胺(EP-PTC)复合膜的形貌及其对以MEH-PPV: EP-PTC复合膜为活性层的太阳电池性能的影响.结果表明：非芳香性溶剂不利于MEH-PPV与EP-PTC的相容，MEH-PPV与EP-PTC两相间形成微米尺寸(0.5—5 μm)的相分离，因而以MEH-PPV: EP-PTC复合膜为活性层的太阳电池中的电荷分离效率较低，进而电池的能量转换效率较低.而 关键词： 太阳电池 相分离 能量转换效率