Development of an immunoassay to detect insect contamination of microalgal products

While the insect fragment count is currently the primary test used for assessing insect contamination of food products, this technique is very problematical for assaying microalgal materials. An account is given of a new immunoassay technique,which is based on an enzyme-linked immunosorbent assay(ELISA) detection of insect myosin and which provides a rapid and convenient means of quantitatively determining the amount of insect contamination in algal product samples with a high degree of replicability. Up to 30 samples can be tested in duplicate in 2.5–3 h. Experiments were carried out with a variety of common contaminant insects of algal products, using various life stages, including Corixidae, Ephydridaeand Chironomidae using both Spirulina (Arthrospira) and Chlorella as typical algal materials. As little as one insect per 50 g sample can readily be detected, with excellent correlation (r² = 0.99) between the number of insects present and the color produced. A matrix analysis to determine the ruggedness of the immunoassay was carried out following the protocols of the AOAC International and established that minor departures in seven variables from the standard assay resulted in no substantial differences. The insect myosin assay offers a quantitative and reliable means for assessing insect contamination of algal materials and should be considered for adoption as a standard method for this type of product. This revised version was published online in August 2006 with corrections to the Cover Date.     

Distribution of SchistoFLRFamide-like immunoreactivity in the adult ventral nervous system of the locust,Schistocerca gregaria

SchistoFLRFamide (PDVDHVFLRF-NH₂) is one of the major endogenous neuropeptides of the FMRF-amide family found in the nervous system of the locust,Schistocerca gregaria. To gain insights into the potential physiological roles of this neuropeptide we have examined the distribution of SchistoFLRFamide-like immunoreactivity in the ventral nervous system of adult locusts by use of a newly developed N-terminally specific antibody. SchistoFLRFamide-like immunoreactivity in the ventral nerve cord is found in a subgroup of the neurones that are immunoreactive to an antiserum raised against bovine pancreatic polypeptide (BPP). In the suboesophageal ganglion three groups of cells stain, including one pair of large posterior ventral cells. These cells are the same size, in the same location in the ganglion and have the same branching pattern as a pair of BPP immunoreactive cells known to innervate the heart and retrocerebral glandular complex of the locust. In the thoracic and abdominal ganglia two and three sets of cells, respectively, stain with both the SchistoFLRFamide and BPP antisera. In the abdominal ganglia the immunoreactive cells project via the median nerves to the intensely immunoreactive neurohaemal organs.     

The effect of plant chemistry on the acceptability of caterpillar prey to the argentine antIridomyrmex humilils (Hymenoptera: Formicidae)

Experiments were performed to test the acceptability of two palatable, cryptic caterpillars, the tobacco hornworm,Manduca sexta, and the cabbage looper,Trichoplusia ni, reared on different diets, to the Argentine ant,Iridomyrmex humilis. Ants preferred larvae reared on artificial diet, groundcherry, or cowpea to tobacco-reared larvae. Ants also preferred larvae reared on artificial diet without nicotine to larvae reared on diet containing nicotine (5% dry wt). Experiments were also performed to test the response of ants to larval extracts and chemicals applied to the surface of palatable prey. Ants did not respond differently to larvae of the potato tuber moth,Phthorimaea operculella, treated with larval extracts or regurgitate from tobacco-reared larvae compared to artificialdiet-reared larvae, but ants were deterred byP. operculella larvae treated with nicotine compared to untreated larvae. The results of this study indicate that caterpillars can derive at least some degree of chemical protection from their food plant without sequestering and storing plant compounds and without the development of elaborate aposematic characteristics.     

Efficient membrane targeting of the chick nicotinic acetylcholine receptor α-subunit in stable insect cell lines

We have synthesised the -subunit of the chick nicotinic acetylcholine receptor (nAChR) in stable, continuous insect (Spodoptera frugiperda) cell lines. A cDNA was integrated randomly into the insect cell genome under control of a baculovius immediate early gene promoter. Transformed cells were obtained by co-transfection of the insect cells with pIEK1.nAChR, encoding the -subunit cDNA, and pIEK1.neo, encoding the neomycin resistance gene. G-418-resistant clones were selected and expanded into continuous cell lines synthesising the chick nAChR -subunit. Using fluorescence microscopy and ligand binding studies we were able to demonstrate efficient membrane targeting of the receptor subunit in the insect cell plasma membrane. Stable insect cell lines may thus have significant advantages over transient baculovirus vectors for the synthesis and characterisation of heterologous receptor proteins.Abbreviations AcNPV Autographa californica nuclear polyhedrosis virus - BTX -bungarotoxin - BSA bovine serum albumin - FITC Fluoroscein isothiocyanate - G418 geneticin-418 - hpi hours post-infection - ie-1 immediate early 1 gene - nAChR nicotinic acetylcholine receptor alpha subunit - Sf Spodoptera frugiperda - tPA tissue plasminogen activator     

Active muscle migration during insect metamorphosis

The R1 abdominal retractor muscles of the insect Tenebrio molitor change position during the course of metamorphosis. These muscles detach from the epidermal tendon cells at their anterior ends, and migrate in a posterior direction, parallel to the body axis, to form completely new attachments shortly before adult emergence. Movement is preceded by the loss of sarcomere structure, and the muscles migrate in a partially dedifferentiated condition, closely accompanied by satellite cells and haemocytes. Movement appears to result from the extension of muscle processes towards the epidermis posterior to the larval attachment sites, which contact reciprocal processes extended from the epidermis. Contacts at the new posterior sites are then reinforced, and relinquished at the anterior. This cycle is subsequently repeated. It is envisaged that migration ceases when the muscles encounter a contour in the epidermal gradient known to specify the position of the adult muscle attachment sites. This positional information may be encoded in the epidermal basal lamina. The muscles then redifferentiate, with concurrent differentiation of new epidermal tendon cells. Development of adult muscle attachments appears to require reciprocal morphogenetic interactions between muscle and epidermis.     

Effects of diet on growth, fecundity and duration of tethered flight of Nezara viridula

The effects of diet during the fifth stadium on performance of adult N. viridula were determined through comparing growth, fecundity, and duration of tethered flight of insects reared on four axenic diets: fresh green bean, raw-shelled peanut, a combination of the two and soaked soybean seed. Females reared on the combination diet were heavier than those reared on green beans and oviposited more than those reared on peanuts. Three-day-old adult females reared on peanuts flew longer than those reared on green beans. More females reared on diets containing mature seed showed a capacity for extended flight and had larger fat bodies at 34-days postemergence. Thus, inadequate diet during the fifth stadium irreversibly affected growth, development, reproduction and flight capacity.

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Résumé Les effets du régime alimentaire pendant le 5ème stade sur les performances imaginales de N. viridula ont été déterminés en comparant la croissance, la fécondité, la durée du vol captif, et l'accumulation de corps gras chez des insectes élevés sur 4 régimes stérilisés. A partir de la formation des larves de 5ème stade, elles ont été alimentées sur haricots verts et arachides, ou transférées sur un régime formé exclusivement d'arachides, ou de haricots verts, ou de graines de soja trempées, cependant la durée du 5ème stade. Les insectes ont été transférés sur un régime de haricots verts et arachides à leur émergence.Les femelles élevées continuellement de haricots verts et arachides étaient plus lourdes (P<0,01) à l'émergence (161,6 mg) que celles élevées sur haricots verts (146,3 mg) pendant le 5ème stade, et pondent plus d'oeufs (P<0,08) pendant une durée de 34 jours (100,9 oeufs) que celles élevées sur arachides (54,7 oeufs). Plus de femelles élevées sur haricots verts et arachides (76,2%) pondent que celles élevées sur arachides (61,1%), soja trempé (62,5%), ou haricots verts (60,9%).Les femelles élevées sur arachides volaient plus longtemps (P < 0,01) (42,9 mn) que celles élevées sur haricots verts (19,8 mn) pendant des vols captifs le 3ème jour après l'émergence. 78% de toutes les femelles élevées sur arachides volaient >-30 mn, contre 50% pour celles élevées sur haricots verts et arachides, ou soja trempé, et 35% pour celles élevées sur haricots verts. Seulement 7% des femelles élevées sur haricots verts volaient à la 60ème minute à la fin du test, contre 25% pour les femelles soja trempé, 33% pour les femelles haricots verts et arachides, et 47% pour celles élevées sur arachides. Les femelles ont été disséquées 34 jours après l'émergence pour déterminer l'importance du corps gras. Les indices de corps gras (Kiritani, 1963) montraient que les femelles élevées sur arachides (1,8) et graines de soja (1,7) ont apparemment un corps gras plus développé que celles élevées sur haricots verts et arachides (1,5), our haricots verts (1,2), pendant le 5ème stade.Ces résultats montrent que l'alimentation pendant le dernier stade larvaire de N. viridula affecte irréversiblement la croissance, la fécondité et l'aptitude au vol des femelles. En estimant que des résultats semblables auraient été obtenus avec des insectes élevés sur ces 4 aliments depuis l'éclosion, haricots verts et arachides constituent le meilleur aliment pour l'élevage de N. viridula. Ce régime fournit une graine mûre, vraisemblablement nécessaire pour l'accumulation d'un corps gras suffisant et une aptitude au vol migratoire, et une plante verte qui fournit les aliments apparement essentiels à l'ovogenèse.

     

Azadirachtin: I-interaction with the development of red cotton bugs

Azadirachtin, a material isolated from Azadirachta indica A. Juss, was applied to various stages of red cotton bugs (Dysdercus koenigii F.). The phytochemical evoked various specific and nonspecific effects during the course of development. Prolonged development, wing deformities unplasticisation of wing lobes, development of wingless adults and larval mortality were the characteristic features.

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Azadirachtine d'Azadirachta indica I: Interaction avec de développement de Dysdercus koenigii

Résumé L'azadirachtine, isolat de Azadirachta indica A. Juss a été utilisée (par traitement du substrat ou par dépôt sur l'insecte) sur un hétérométabole, Dysdercus koenigii F. Cette substance phytochimique provoque divers effets, spécifiques ou non, pendant le développement.Les modifications caractéristiques sont: un développement prolongé, de déformations alaires, une absence de plastification des bourgeons alaires, une production d'adultes sans ailes et une mortalité nymphale.De tels résultats suggèrent des études complémentaires pour mettre en évidence une hormone d'éclosion chez D. koenigii avec laquelle interférerait l'azadirachtine.

     

Gypsy moth cell lines divergent in viral susceptibility

Summary A series of cell lines unique in insect virus susceptibility pattern have been isolated from the ovaries of the gypsy moth (Lymantria dispar: Lepidoptera: Lymantriidae) on a synthetic medium with mammalian and avian serum supplementation. Growth curves showed the poorest growth occurring on peptone-based media with somewhat better growth on amino-acid-based media. The best growth was obtained with combined media. Serological study distinguished the present cell lines from one another and from cell lines derived from other insect species grown routinely in the same laboratory. Baculovirus susceptibility among the new lines varied from no response to a specific complete replication response upon challenge by the homologous (gypsy moth) nuclear polyhedrosis virus. This research was funded in part through a reimbursable agreement with the U.S. Forest Service.     

Cell cycle analysis and synchronization of the TN-368 insect cell line

Summary A cell cycle analysis of theTrichoplusia ni (TN-368) insect cell line is described. By means of autoradiography and percent labeled metaphase data, the cell cycle parameters were determined to be as follows: S, 4.5 hr; G₂, 8.5 hr; M, 0.5 hr; G₁, 1.0 hr; the total cell time being 14.5 hr. A synchronization procedure using 50mm thymidine in a double block procedure was used to provide a method of obtaining a large number of cells in particular cell cycle phases, especially S and G₂. This work was supported in part by U.S. Environmental Protection Agency Grant R-802516.     

Plant species diversity and ecological districts of the Sinai desert

Summary Lists of vascular plant species for each of the 12 ecological districts of Sinai were prepared. In all 78,000 observations on altogether 812 species were analyzed. The number of districts where each species occurred was also counted. Linear regressions were calculated for the log-transformed values of species/area, species/altitude and multiple regression of species on both area and altitude.The regression of species on area for the 12 ecological districts of Sinai gave the equation: log S = 1.0309+0.3 log A The value of z=0.3 is higher than 0.222—the measured overall value for the continents and higher than the values for the Sahara, California mainland, the British Isles, and the Netherlands. However, it is lower than the values for the Galápagos Archipelago and the California Islands.The number of species supported in districts characterized by fissured limestone, gravel plains, chalk, marl, sandstones, sands or fissured magmatic and metamorphic rocks is close to the regression of species on area, whereas large outcrops of smooth-faced rocks are relatively richer in total number of species as well as in stenotople species.It is suggested that the high gamma diversity of Sinai as compared with other parts of the world is primarily due to its environmental heterogeneity. Sinai being a meeting place of three phytogeographical regions and to past climatic changes. The effect of smooth-faced rock outcrops as conducive to providing refugia must also be taken into account.This work is based on a comprehensive study of the vegetation of Sinai directed by Prof. G. Orshan, with the collaboration of Dr. A. Shmida, the present author and the late Prof. N.H. Tadmor and Dr. G. Halevy.