Cadmium effect on growth and biochemical traits of Gypsy moth larvae (Lymantria dispar L.)

Gubar (Lymantria dispar L.) spada u red najinvazivnijih polifagnih vrsta insekata, koja naseljava šume, voćnjake i urbana staništa. U ovom radu je kod larvi gubara trećeg dana četvrtog larvenog stupnja, ispitivan akutni i hroničan uticaj dve koncentracije kadmijuma (10 i 30 μgCd/g suve hrane), kao i oporavak od istih, na različite osobine larvi: komponente adaptivne vrednosti (mortalitet, trajanje prvog stupnja, trajanje razvića do ulaska u četvrti larveni stupanj, masa i relativna brzina rasta larvi), aktivnosti enzima srednjeg creva (proteaza, tripsin, leucin aminopeptidaza, α i β-glikozidaza, esteraza i glutation-S-transferaza), kao i procenat mase srednjeg creva u odnosu na masu larve. Kod svih ispitivanih enzima, osim kod GSTa, prisutan je trend pada aktivnosti na kadmijumu u odnosu na kontrolu. Hronična izloženost kadmijumu nije pokazala značajan uticaj na mortalitet prvog stupnja, kao i ukupan mortalitet, masa larvi se smanjuje pri dugotrajnim tretmanima, dok se u toku akutnog tretmana brzina rasta larvi povećava. Analize varijanse su pokazale da se kod proteolitičkih enzima aktivnost menja u zavisnosti od koncentracije metala. Između proteolitičkih enzima na svim tretmanima najverovatnije postoji neka vrsta zajedničke regulacije sinteze ili sekrecije u prisustvu kadmijuma. Regulacioni mehanizmi β-glikozidaze i GSTa se razlikuju u odnosu na ostale proučavane enzime. Zbog ograničenih energetskih resursa postoji veliki broj negativnih korelacija između mase i enzima. Znak korelacija između α i β-glikozidaza se menja na stresu u odnosu na kontrolu. Na različitim tretmanima kadmijumom plastičnost enzima se menja. Pošto se indeks fenotipske plastičnosti ne menja u toku delovanja kadmijuma, promena osobina adaptivne vrednosti ili njihova plastičnost, se ne mogu uzeti kao dobri indikatori prisustva kadmijuma. Zbog velike osetljivosti leucin aminopeptidaze i esteraze u toku akutnog stresa, postojanja korelacija sa velikim brojem digestivnih enzima, velike plastičnosti odgovora u stresnim uslovima, ova dva enzima bi se, uz dalja istraživanja, mogla uzeti kao potencijalni biomarkeri na prisustvo kadmijuma.The Gypsy moth (Lymantria dispar L.) represents one of the most invasive polyphagous insect species, which inhabits forests, orchards and even urban habitats. We investigated the acute and chronic effects of two cadmium concentrations (10 and 30 μgCd/g dry food), as well as the recovery from chronic stress on various traits of 4th instar gypsy moth larvae: fitness components (mortality, duration of development of the first larval instar, duration of development from hatching to molting into the 4th instar, larval mass and relative growth rate), midgut enzyme activities (activity of protease, trypsin, leucine aminopeptidase, α and β-glucosidase, esterase and glutathione S-transferase), as well as the percentage of the midgut mass in relation to the larval mass. All examined enzymes, with the exception of the glutathione S-transferase, showed a decrease of activity in relation to control. Larval mass decreased during the long-term treatment, while the acute treatment resulted in the growth rate increase. The analysis of variance showed that the activity of proteolytic enzymes depended on metal concentration. Significant broad-sense heritability was discovered after short-term exposure to cadmium. Regulation mechanisms of β-glucosidase and glutathione S-transferase differ in relation to the rest of the examined enzymes. There are a large number of negative correlations between larval mass and enzymes due to the limited energy resources. The correlation sign between α and β- glucosidases changes during the stress conditions in relation to the control. During different cadmium treatments plasticity of enzymes is changed. Changes of life history traits or their plasticity cannot be good indicators of cadmium presence. Due to a great sensitivity of leucine aminopeptidase and esterase during the acute stress, correlated response to a large number of digestive enzymes, great plasticity in stressful conditions, these two enzymes, with further research, could be potential biomarkers for cadmium presence

Biogenic amines in protocerebral A2 neurosecretory neurons of Lymantria dispar L. (Lepidoptera:Lymantriidae): Response to trophic stress

The number, morphometric parameters and amount of aminergic neurosecretory product of protocerebral A2 neurosecretory neurons were investigated in the fifth instar of Lymantria dispar caterpillars, following a suitable or unsuitable trophic regime. Caterpillars originated from two populations (Quercus rubra or Robinia pseudoacacia forest) and were differently adapted to trophic stress, i.e. feeding on locust tree leaves - unsuitable host plant. The number of neurosecretory neurons was higher in the caterpillars originated from Robinia population than in Quercus population, regardless of feeding. A2 neurosecretory neurons, nuclei and their nucleoli were larger in caterpillars fed with unsuitable leaves in both populations. There was more aminergic product in the A2 neurosecretory neurons of the caterpillars fed with unsuitable leaves independently of population origin

Biogenic amines in protocerebral A2 neurosecretory neurons of Lymantria dispar L. (Lepidoptera:Lymantriidae): Response to trophic stress

The number, morphometric parameters and amount of aminergic neurosecretory product of protocerebral A2 neurosecretory neurons were investigated in the fifth instar of Lymantria dispar caterpillars, following a suitable or unsuitable trophic regime. Caterpillars originated from two populations (Quercus rubra or Robinia pseudoacacia forest) and were differently adapted to trophic stress, i.e. feeding on locust tree leaves - unsuitable host plant. The number of neurosecretory neurons was higher in the caterpillars originated from Robinia population than in Quercus population, regardless of feeding. A2 neurosecretory neurons, nuclei and their nucleoli were larger in caterpillars fed with unsuitable leaves in both populations. There was more aminergic product in the A2 neurosecretory neurons of the caterpillars fed with unsuitable leaves independently of population origin.Projekat ministarstva br. 17302

Activity of gypsy moth dorsolateral neurosecretory neurons under increased rearing density

Lymantria dispar caterpillars were reared under two different rearing densities for the first three days of the 4th larval instar: 5 larvae that were kept in a Petri dish (V = 80 ml) belonged to the intense stress (D1 group); 5 larvae that were kept in a plastic cup (V = 300ml) belonged to the group exposed to less intense stress (D2 group). In the control group, single larvae were reared in a Petri dish. Morphometric changes in L1, L2 and L2’ dorsolateral neurosecretory neurons (nsn) were analyzed. After keeping 5 larvae in a Petri dish, the size of L2 neurosecretory neurons (nsn) significantly increased. Rearing 5 larvae in a plastic cup significantly increased the size of L1 nsn nuclei and the number of L2’nsn. A decrease in relative band densities in the region of molecular masses (11-15 kD) that correspond to prothoracicotropic hormones in the gypsy moth was observed in the electrophoretic profiles that were obtained after both treatments in comparison to the control group. [Acknowledgments. This study was supported by the Serbian Ministry of Education and Science (Grant No. 173027).

Morphometric changes of corpora allata in Morimus funereus müls. (Cerambycidae) larvae during thermal stress

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Sensitivity of gypsy moth neurosecretory neurons to acute thermal stress

In gypsy moth caterpillars exposed to a temperature of 35°C (for 1, 12 and 24 h and caterpillars that were exposed to elevated temperature for 12 h and were allowed to recover for 12 h at 23°C), changes in the brain protein profiles and morphometric characteristics of A1’ medial and L2 lateral protocerebral neurosecretory neurons were analyzed. In all groups, protein bands with a molecular mass corresponding to that of members of heat-shock protein families were detected, indicating that acute exposure to this temperature likely induced the synthesis of HSP. Increased morphometric parameters of A1’ neurons and the large amount of neurosecretory material in the neuron body implicate that the temperature of 35°C is not in the temperature range that exerts stimulatory effects on growth and survival. Changes in the morphometric characteristics of L2 neurosecretory neurons from the lateral part of the protocerebrum, and retention of neurosecretory material in their cytoplasm indicate a low level of secretion

Effects of tannic acid on trypsin and leucine aminopeptidase activities in gypsy moth larval midgut

The effects of allelochemical stress on genetic variations in the specific activities of gypsy moth digestive enzymes (trypsin and leucine aminopeptidase) and relative midgut mass (indirect measure of food consumption), as well as variability in their plasticity, were investigated in fifth instar gypsy moths originating from two populations with different trophic adaptations (oak and locust-tree forests). Thirty-two full-sib families from the Quercus population and twenty-six full-sib families from the Robinia population were reared on an artificial diet with or without supplementation with tannic acid. Between population differences were observed as higher average specific activity of trypsin and relative midgut mass in larvae from the Robinia population. Significant broad-sense heritabilities were observed for the specific activity of trypsin in the control state, and for specific activity of leucine aminopeptidase in a stressful environment. Significantly lower heritability for relative midgut mass was recorded in larvae from the Robinia population reared under stressful conditions. Significant variability of trypsin plasticity in larvae from both populations and significant variability of leucine aminopeptidase plasticity in larvae from the Robinia population point to the potential for the evolution of enzyme adaptive plastic responses to the presence of stressor. Non-significant across-environment genetic correlations do not represent a constraint for the evolution of enzyme plasticity. [Projekat Ministarstva nauke Republike Srbije, br. 173027

Changes in Lymantria dispar protocerebral neurosecretory neurons after exposure to cadmium

Gypsy moth 4th instar caterpillars were fed for 3 days with an artificial diet supplemented with increasing cadmium (Cd) concentrations (0, 10, 30, 100 and 250 μg⁄g of dry food weight). Changes in the morphometric characteristics of A1' dorso-medial and L2 dorso-lateral neurosecretory neurons (nsn) were analyzed. In the A1' nsn, Cd supplements led to an enhanced nuclear size, except in the group treated with 250 μg Cd⁄g in the form of dry food. The size of L2 type nsn was increased in the groups provided with 30 and 100 μg Cd⁄g, while no differences in the size of nuclei was detected in L2 neurons among the experimental groups.Projekat ministarstva br. 7302

Benzo[a]pyrene-induced changes in carboxylesterase, acetylcholinesterase and heat shock protein 70 of Lymantria dispar (Lepidoptera: Lymantriidae) from unpolluted and polluted forests

Plant vegetation accumulates polycyclic aromatic hydrocarbons (PAHs) among which benzo[a]pyrene (B[a]P) is recognized as being very toxic, including cancerogenic. Lymantria dispar L. larvae are sensitive to changes in the environment, providing potential signs of pollutant presence. We examined the chronic effects of two concentrations of B[a] P on the activity of carboxylesterase (CaE), acetylcholinesterase (AChE) and heat shock protein 70 (Hsp70) levels in the brain tissue of two populations of L. dispar larvae, originating from unpolluted and polluted habitats. We found that the relative growth rate was significantly lower in both populations and that only larvae from polluted forests were sensitive to low B[a]P concentrations, exhibiting a significant increase in brain tissue CaE activity and Hsp70 concentration. AChE activity showed no changes in response to B[a]P exposure in either population. Examined biochemical parameters indicate that their sensitivity to chronic treatment with B[a]P was highly dependent on the pre-exposure history of L. dispar larvae, suggesting that they could be promising biomarkers of B[a]P and PAH pollution in forest ecosystem

Effects of tannic acid on trypsin and leucine aminopeptidase activities in gypsy moth larval midgut

The effects of allelochemical stress on genetic variations in the specific activities of gypsy moth digestive enzymes (trypsin and leucine aminopeptidase) and relative midgut mass (indirect measure of food consumption), as well as variability in their plasticity, were investigated in fifth instar gypsy moths originating from two populations with different trophic adaptations (oak and locust-tree forests). Thirty-two full-sib families from the Quercus population and twenty-six full-sib families from the Robinia population were reared on an artificial diet with or without supplementation with tannic acid. Between population differences were observed as higher average specific activity of trypsin and relative midgut mass in larvae from the Robinia population. Significant broad-sense heritabilities were observed for the specific activity of trypsin in the control state, and for specific activity of leucine aminopeptidase in a stressful environment. Significantly lower heritability for relative midgut mass was recorded in larvae from the Robinia population reared under stressful conditions. Significant variability of trypsin plasticity in larvae from both populations and significant variability of leucine aminopeptidase plasticity in larvae from the Robinia population point to the potential for the evolution of enzyme adaptive plastic responses to the presence of stressor. Non-significant across-environment genetic correlations do not represent a constraint for the evolution of enzyme plasticity.Projekat ministarstva br. 17302