Biochemical efficacy, molecular docking and inhibitory effect of 2, 3-dimethylmaleic anhydride on insect acetylcholinesterase.

Evolution of resistance among insects to action of pesticides has led to the discovery of several insecticides (neonicotinoids and organophosphates) with new targets in insect nervous system. Present study evaluates the mode of inhibition of acetylchlonesterase (AChE), biochemical efficacy, and molecular docking of 2,3-dimethylmaleic anhydride, against Periplaneta americana and Sitophilus oryzae. The knockdown activity of 2,3-dimethylmaleic anhydride was associated with in vivo inhibition of AChE. At KD99 dosage, the 2,3-dimethylmaleic anhydride showed more than 90% inhibition of AChE activity in test insects. A significant impairment in antioxidant system was observed, characterized by alteration in superoxide dismutase and catalase activities along with increase in reduced glutathione levels. Computational docking programs provided insights in to the possible interaction between 2,3-dimethylmaleic anhydride and AChE of P. americana. Our study reveals that 2,3-dimethylmaeic anhydride elicits toxicity in S. oryzae and P. americana primarily by AChE inhibition along with oxidative stress.

Desenvolvimento de Periplaneta australasiae Fabricius (Blattodea, Blattidae) em temperatura constante de 30º c e em temperatura ambiente de laboratório / Development of Periplaneta australasiae Fabricius (Blattodea, Blattidae) at a constant temperature of 30° c and at ambient laboratory temperature

Objetivou-se avaliar o desenvolvimento de ovos e ninfas de Periplaneta australasiae (Fabricius) à temperatura de 30 ± 0,2° C, umidade relativa 80 ± 15% e fotofase de 12h e em condições ambientais de laboratório, sem controle de temperatura e umidade relativa; visando à subsídios para medidas de prevenção e controle. As ootecas foram individualizadas em tubos de ensaio até a eclosão. As ninfas foram transferidas para cubas de vidro e alimentadas com ração comercial para coelhos e água ad libitum até a emergência das imagos. Avaliou-se, período de incubação, número de ovos/ ooteca, viabilidade de ovos, número de ninfas/ooteca, período ninfal, viabilidade de ninfas e período ovo/adulto. A diferença do período médio de incubação à temperatura de 30° C (38 dias) e no ambiente (44,5 dias) foi significativa (p < 0,0001); eclodiram, em média 18,1 ninfas/ooteca a 30° C e 21 ninfas/ooteca em condições ambientais (p = 0,006); o período médio de ninfa a 30° C foi de 155,9 dias e no ambiente 279,7 dias (p < 0,0001); a viabilidade de ninfas foi superior a 50%, tanto a 30° C (55,1%) quanto em condições de laboratório (57,2%); no período médio de ovo-adulto de P. australasiae, houve diferença significativa (p< 0,001) entre a temperatura de 30 °C (194,1 dias) e em condições ambientais de laboratório (337,3 dias). Em condições de laboratório, os períodos de incubação, de ninfa e de ovo-adulto de P. australasiae foram aumentados em relação à temperatura de 30° C, não ocorrendo, entretanto, perda nem redução de viabilidade em nenhuma das fases.

With the objective of obtaining standards of measurement for prevention and control, this study compared the development of the eggs and nymphs of Periplaneta australasiae (Fabricius) at a temperature of 30 ± 0.2° C, relative humidity 80 ± 15% and photoperiod of 12 hours versus ambient conditions in the laboratory without controls of temperature and RH. Single ootheca were maintained in test tubes until ecolosion, and nymphs were transferred to glass cubes and fed commercial rabbit ration and water ad libitum until emergence of the imagos. The incubation period, number of eggs/ ootheca, viability of the eggs, number of nymphs/ootheca, nymphal duration, viability of nymphs and duration of egg to adult were all evaluated. The mean difference in the incubation period between the temperature of 30° C (38 days) and ambient conditions (44.5 days) was significant (p < 0.0001); a mean of 18.1 nymphs/ootheca ecloded at 30° C, while 21 nymphs/ootheca ecloded under ambient conditions (p = 0.006); the mean nymphal period at 30° C was 155.9 days while for the ambient it was 279.7 days (p < 0.0001); nymphal viability was greater than 50% for both the 30° C laboratory (55.1%) and the ambient (57.2%); and the mean period from egg to adult of P. australasiae was significantly different (p < 0.001) between the 30° C temperature (194.1 days) and the ambient conditions of the laboratory (337.3 days). Under ambient conditions, the duration of nymphal incubation and egg to adult development of P. australasiae were increased relative to the temperature of 30° C without a reduction in viability in any of the stages.

[Effects of several exorphins and endorphins on the escape reaction of the cockroach Periplaneta americana under elevated temperature conditions].

The ability of several alimentary opioid peptides (exorphin C, rubiscolin-5, cytochrophi-4) and endorphins (met-enkephalin, dynotphin A(1-10), beta-neoendorphin) to change the escape reaction of the cockroaches Periplaneta americana at their placement into a hot chamber was studied. The ED50 values increasing twice the insect stay time in the hot chamber as well as duration and dynamics of the effects were determined. It has been shown that ED5 decreases statistically significantly with increase of the length of the peptide molecule and its affinity of duration of the effects and to an increase of their affinity to delta-receptors - to prolongation of the reaction (more than150 min). In the group of alimentary peptides (exorphins) the most active was a fragment of D-ribulose-1,5-biphosphate carboxylase/oxigenase rubiscolin-5 (ED5 = 386 nM per individual). This might indicate a specific ability of some plant proteins to regulate (decrease) the insect protective behavior.

Proton-pyrophosphatase and polyphosphate in acidocalcisome-like vesicles from oocytes and eggs of Periplaneta americana.

Acidocalcisomes are acidic organelles containing large amounts of polyphosphate (poly P), a number of cations, and a variety of cation pumps in their limiting membrane. The vacuolar proton-pyrophosphatase (V-H(+)-PPase), a unique electrogenic proton-pump that couples pyrophosphate (PPi) hydrolysis to the active transport of protons across membranes, is commonly present in membranes of acidocalcisomes. In the course of insect oogenesis, a large amount of yolk protein is incorporated by the oocytes and stored in organelles called yolk granules (YGs). During embryogenesis, the content of these granules is degraded by acid hydrolases. These enzymes are activated by the acidification of the YG by a mechanism that is mediated by proton-pumps present in their membranes. In this work, we describe an H(+)-PPase activity in membrane fractions of oocytes and eggs of the domestic cockroach Periplaneta americana. The enzyme activity was optimum at pH around 7.0, and was dependent on Mg(2+) and inhibited by NaF, as well as by IDP and Ca(2+). Immunolocalization of the yolk preparation using antibodies against a conserved sequence of V-H(+)-PPases showed labeling of small vesicles, which also showed the presence of high concentrations of phosphorus, calcium and other elements, as revealed by electron probe X-ray microanalysis. In addition, poly P content was detected in ovaries and eggs and localized inside the yolk granules and the small vesicles. Altogether, our results provide evidence that numerous small vesicles of the eggs of P. americana present acidocalcisome-like characteristics. In addition, the possible role of these organelles during embryogenesis of this insect is discussed.

Genome size of termites (Insecta, Dictyoptera, Isoptera) and wood roaches (Insecta, Dictyoptera, Cryptocercidae).

The evolution of genome size has been discussed in relation to the evolution of various biological traits. In the present study, the genome sizes of 22 dictyopteran species were estimated by Feulgen image analysis densitometry and 6-diamidino-2-phenylindole (DAPI)-based flow cytometry. The haploid genome sizes (C-values) of termites (Isoptera) ranged from 0.58 to 1.90 pg, and those of Cryptocercus wood roaches (Cryptocercidae) were 1.16 to 1.32 pg. Compared to known values of other cockroaches (Blattaria) and mantids (Mantodea), these values are low. A relatively small genome size appears to be a (syn)apomorphy of Isoptera + Cryptocercus, together with their sociality. In some phylogenetic groups, genome size evolution is thought to be influenced by selective pressure on a particular trait, such as cell size or rate of development. The present results raise the possibility that genome size is influenced by selective pressures on traits associated with the evolution of sociality.

Progesterone in Periplaneta americana and Neobellieria bullata adults from the procuticle phase until first progeny production.

A significant amount of progesterone-like immunoreactive material (150 ng/g) was measured by EIA in the procuticle phase of adult of both sexes of Periplaneta americana. This peak markedly decreased to 1-10 ng/g during sclerotization and was unlikely to be of dietary origin. In the case of 0-hr-old P. americana adults 96-98% of progesterone-like material was localized in the digestive tract and Malpighian tubules. In contrast, a relatively low level of progesterone-like immunoreactive material was measured in 0-hr-old Neobellieria bullata adults. Activity of 3beta-HSD/isomerase converting pregnenolone to progesterone was high (22-43 fmol/mg protein/20 min) in 0-hr-old P. americana adults and significantly fell during sclerotization. High progesterone levels (13-16 ng/g), measured by HPLC-RIA, coexist with high levels of 3beta-HSD/isomerase activity. Orally active human contraceptives (ethisterone, ethynodiol, ethynodiol diacetate, lynestrenol, mestranol, norgestrel, norethynodrel, tamoxifen citrate, and mifepristone) which act on mammalian steroid receptors had no significant effects on progeny production in either polytrophic or meroistic insect ovaries even at concentration of 5000 mg/kg.

The effect of temperature on flight initiation in the cockroach Periplaneta americana.

The effect of temperature during development of the flight system of the American cockroach was tested. Animals were raised at three different temperatures from egg cases to adulthood then various parameters of flight were tested. Animals raised at cooler temperatures were found to require significantly stronger wind puffs to initiate flight than did animals raised in warmer temperatures. Some animals were raised at one temperature during embryonic development (i.e. while the animals were still in egg cases) and then switched to a different temperature for nymphal development. In these cases only the temperature experienced during embryonic development had a significant permanent effect on flight. Parameters associated with pattern generation such as interburst interval were not affected by temperature during either developmental period.

Preservation of motoneuron electrotonic characteristics during postembryonic growth.

The postembryonic developmental period of the nervous system involves, among other processes, an increase in the dimensions of individual neurons. In the present study we examine whether the characteristic electronic parameters that underlie the integrative properties of the neuron (L, the electrotonic length; rho, dendritic to somatic conductance ratio; and tau m, the membrane time constant) are established before, during, or after the neurons reach their final adult size. A second aspect that we analyze are the mechanisms by which the neurons adjust their electrotonic parameters during the growth period. The dimensions of the soma and dendrites of the phallic motoneurons (Ph.m.n.) of the cockroach, Periplaneta americana, increase during the last postembryonic developmental stages and metamorphosis to adult by a factor of 1.75 and 1.5, respectively. This increase is not associated with major changes in the morphological outline of the neuron but is associated with a significant decrease in the input resistance (Rin) by a factor of 6 (from 120 to 20 M omega). The dendritic to somatic conductance ratio (rho) is maintained constant. Morphological analysis of cobalt-filled neurons reveals that the neurons can be represented by an equivalent cylinder (Rall, 1969). Calculations of the electrotonic length (L) of the equivalent cylinder from measurements of the time constants tau m and tau 1 (Rall, 1969) revealed that L is constant throughout the period of neuronal growth (1.29 +/- 0.24 lambda in adult, and 1.13 +/- 0.28 lambda in nymph). The mechanism by which the electrotonic parameters are maintained in spite of growth is continuous adjustment of the diameter and length of the various neuronal segments, rather than by changes in the biophysical properties of the neuron. Several implications in relation to mechanisms of integration and input-output relations during growth and development of neurons are discussed in view of our findings.

Ontogeny of the invertebrate humoral immune response: studies on various developmental stages of the American cockroach (Periplaneta americana).

Earlier studies revealed that a specific adaptive humoral immune response can be induced in the American cockroach (Periplaneta americana) to the soluble protein complex, Honeybee venom (HBT). We have undertaken a series of ontogenetic studies to determine if there are differences between the protective responses of roaches representing different developmental stages. Our results indicated that the response to HBT in the immature cockroach (nymphs weighing 200-500 mg) was characterized by a significant lag period before immune protection began to develop. However, by the second week of the response, reactivity was comparable to that of the adult. Old adult male roaches (animals 5 months into adulthood) displayed a significant decline in reactivity during the early phases of the response in comparison to younger adults, and in general, appeared to be less vigorous in generating protection. Both the nymph and old adult roaches demonstrated good secondary responsiveness. The results from these experiments indicated that the developmental stage of the roach could be directly correlated to the degree of immunocompetency possessed by the animal. This is similar to the ontogenetic sequence typically found for immune reactivity in vertebrates.

Ontogeny of endocrine cells in the gut of the insect Periplaneta americana.

The ontogeny of the endocrine cells of the gut of the cockroach Periplaneta americana was studied by immunohistochemistry. During embryogenesis, the midgut begins to be formed as an outgrowth of the foregut and hindgut invaginations. Gut endocrine cells with pancreatic polypeptide (PP)-like immunoreactivity begin to appear at the anterior and posterior ends of the forming midgut. These cells are restricted to the midgut epithelium, and no mitotic cells with PP-like immunoreactivity are observed. These results strongly suggest that the gut endocrine cells, at least those with PP-like immunoreactivity, are derived from precursor cells they have in common with other epithelial cells of the midgut.

Formation of the retina-lamina projection of the cockroach: no evidence for neuronal specificity.

There is a topographical mapping of neural elements onto the lamina neuropile of the optic lobe of the cockroach, such that adjacent ommatidia project to adjacent points (optic cartridges) in the lamina neuropile. Postembryonic growth of the compound eye occurs by addition of new ommatidia to its growing margin. Retinula axons grow from the newly formed ommatidia to the lamina. By transplantation experiments in which the position or the orientation of retinal material is altered, it is shown that retinula axons do not make connections in the lamina with respect to their old position and orientation, but rather, in keeping with their new situations, apparently maintaining a retinotopic mapping upon the optic lobe.

Postembryonic growth of the compound eye of the cockroach.

The postembryonic growth of the compound eye of the cockroach Periplaneta americana involves increases in the size of the individual ommatidia as well as a 35-fold increase in the number of ommatidia. These ommatidia are added to the anterior, dorsal, and ventral margins of the eye by means of an almost continuous process of cell division in the proliferation zone in these margins. This proliferation phase is followed by a process of maturation of bundles of 'pre-ommatidial' cells into mature ommatidia, a process which involves further cell division. Processes involved in compound-eye development are investigated by eye margin grafting, histological techniques and cell proliferation studies.

Postembryonic development of the antennal lobes in Periplaneta americana L.

The postembryonic development of the antennal lobes of Periplaneta americana L. was examined with light- and electron-microscopical methods. There is no difference in the number of glomeruli and neurons in the antennal lobes of larval and adult animals. At hatching, the first larva already possesses the adult number of approximately 125 glomeruli and 500 to 560 deutocerebral neurons in the dorsolateral cell group of each antennal lobe. During postembryonic development the volume of the deutocerebral neurons increases three- to fourfold. The glomeruli of the first larva have about 7% of the volume of the corresponding adult glomeruli. Since number, pattern, and size ratio of glomeruli (with the exception of the macroglomerulus) are constant in all larval stages and adult animals, it is possible to identify individual glomeruli. During the whole postembryonic development the ordinary glomeruli show a continuous volume increase, which parallels the increase in antennal sensory input. The macroglomerulus develops by way of special growth of two to four neuropil units, but not before the last three to four larval stages and only in males. Its growth precedes the formation of antennal pheromone receptors during the final molt; these receptors are known to project into the macroglomerulus. The development of the macroglomerulus in the last larval stages of the male may be caused by a genetically fixed growth program of specific deutocerebral neurons.

The eye margin and compound-eye development in the cockroach: evidence against recruitment.

The compound eye of the cockroach nymph grows from stadium to stadium by the addition of new ommatidia to the growing edge of the eye. By a series of transplant operations on Periplaneta americana and from SEM studies on Gromphadorhina portentosa it is shown that the proliferating region of the eye margin is a budding zone. There is no recruitment of larval head-capsule epidermis into the eye.

The influence of denervation of the maturation of the flight muscle of Periplaneta americana.

The metathoracic musculature of the American cockroach Periplaneta americana was denervated by dissecting the nerves originating in the metathoracic ganglion on one side within 2 days after the last moult. The biochemical and morphological differences between normal and denervated musculature were followed two weeks later. Denervation completely prevents the gain in weight, protein content and cytochrome oxidase activity which occur in normal muscles during the two weeks after previous moulting. The content of phosphoarginine in the denervated muscle does not differ from the control muscle and is lower than in young muscles. The ability to resynthesize phosphoarginine postanaerobically is substantially lower in the denervated muscle than in control muscles, or in young muscles immediately after the last moult. The ultramicroscopic structure of the denervated muscle differs from that of the normal muscle: the mitochondria possess less cristae, the matrix between them appears less dense and the sarcoplasmatic reticulum is less developed. Thus the denervated muscle not only fails to mature, but also developes serious structural and biochemical disorders.