Instar Larval Stage Research Articles

Effects of short‐term heat stress on the growth and development of Bradysia cellarum and Bradysia impatiens

AbstractBradysia cellarum Frey and Bradysia impatiens Johansen are major pests of vegetable crops, as well as edible mushrooms and ornamental plants, and damage to hosts resulting in economic losses. Temperatures above the optimum levels for these pests have been predicted to regulate their population growth during summer. The aim of the present study was to examine the effects of both heat stress and exposure time on the growth and development of eggs, larvae and pupae for two Bradysia species. The egg stage, egg hatching rate, 4th instar larval stage, pupation rate, pupal stage and adult emergence rate were observed after exposing at high temperatures of 34°C, 37°C and 40°C for 1, 2, 4 and 6 hr. The results showed that 34°C, 37°C and 40°C for 1‐, 2‐, 4‐ and 6‐hr exposure treatments prolonged the developmental stage of egg, 4th instar larva and pupa, while decreasing the egg hatching rate, pupation rate and adult emergence rate. This suggests that increasing temperature or prolonging exposure time to the heat stress could significantly affect insect survival, growth and development. Our study could provide an ecological basis for pests’ management strategy by using short‐term heat stress.

Lar maintains the homeostasis of the hematopoietic organ in Drosophila by regulating insulin signaling in the niche.

Stem cell compartments in metazoa get regulated by systemic factors as well as local stem cell niche-derived factors. However, the mechanisms by which systemic signals integrate with local factors in maintaining tissue homeostasis remain unclear. Employing the Drosophila lymph gland, which harbors differentiated blood cells, and stem-like progenitor cells and their niche, we demonstrate how a systemic signal interacts and harmonizes with local factor/s to achieve cell type-specific tissue homeostasis. Our genetic analyses uncovered a novel function of Lar, a receptor protein tyrosine phosphatase. Niche-specific loss of Lar leads to upregulated insulin signaling, causing increased niche cell proliferation and ectopic progenitor differentiation. Insulin signaling assayed by PI3K activation is downregulated after the second instar larval stage, a time point that coincides with the appearance of Lar in the hematopoietic niche. We further demonstrate that Lar physically associates with InR and serves as a negative regulator for insulin signaling in the Drosophila larval hematopoietic niche. Whether Lar serves as a localized invariable negative regulator of systemic signals such as insulin in other stem cell niches remains to be explored.

Characterisation of GST genes from the Hyphantria cunea and their response to the oxidative stress caused by the infection of Hyphantria cunea nucleopolyhedrovirus (HcNPV)

The fall webworm, Hyphantria cunea (Drury) (Lepidoptera: Noctuidae), is a major pest found in forests. In this study, the effects of Hyphantria cunea nucleopolyhedrovirus (HcNPV) infection on the transcription levels and activities of glutathione S-transferases (GSTs) in H. cunea were determined. In the present study, 18 GST family genes were identified from the H. cunea transcriptome dataset by using bioinformatic analyses. These GST genes were classified into cytosolic (15 genes) and microsomal (three genes) classes. The 15 cytosolic GST genes belonged to four different subclasses (epsilon, sigma and delta). The all GST genes, especially GSTe4, showed high expression levels in egg and 1st~4th instar larval stage while their low expression levels in 5th~7th instar larvae using real-time quantitative PCR analysis. However, the expression levels of the 18 GST genes were varied after exposure to sublethal doses of HcNPV. The expression levels of most GSTs were downregulated and upregulated at low and high concentrations of HcNPV, respectively. The corresponding total GST activities also showed similar patterns. In H. cunea, changes in the expression levels and enzymatic activities of GSTs after exposure to HcNPV indicated that they may have important functions in the defense against HcNPV, and the stress, which may be reflected by the high GST enzymatic activities.

Kunitz-type protease inhibitor BmSPI51 plays an antifungal role in the silkworm cocoon

The protease inhibitors found in silkworm cocoons can be divided into several families, a majority of which contain serpin, TIL, or Kunitz domains. Previously, it has been reported that TIL-type protease inhibitors have antimicrobial activity. To date, however, it has not been determined whether the Kunitz-type protease inhibitor BmSPI51, the most abundant of cocoon protease inhibitors, plays an antimicrobial role. Thus, in this study, we sought to determine the biological role of BmSPI51 in silkworm cocoons. Our results obtained from real-time quantitative reverse transcription PCR and immunofluorescence analyses indicate that BmSPI51 is expressed exclusively in the silk glands during the larval fifth instar stage and is subsequently secreted into cocoon silk. Moreover, at a molar ratio of 1:1, BmSPI51 produced via prokaryotic expression exhibited inhibitory activity against trypsin and also proved to be highly stable over wide ranges of temperature and pH values. The expression of BmSPI51 was also found to be significantly upregulated in the larval fat body after infection with three species of fungi, namely, Candida albicans, Beauveria bassiana, and Saccharomyces cerevisiae. In vitro inhibition tests revealed that BmSPI51 significantly inhibited the sporular growth of all three of these fungal species. Further, results obtained from a binding assay showed that BmSPI51 binds to β-d-glucan and mannan on the surface of fungal cells. In this study, we, thus, revealed the antimicrobial activity of BmSPI51 and its underlying mechanism in silkworm, thereby contributing to our present understanding of defense mechanisms in silkworm cocoons.

Purification and characterization of protease inhibitors from rice bean (Vigna umbellata) and their efficacy against Spodoptera litura (Fabricius) gut proteinases

A protease inhibitor from rice bean was isolated and purified to assess its potential against gut proteases of Spodoptera litura, a devastating pest of several economically important crops globally. The purified inhibitor showed a single band protein on SDS-PAGE corresponding to molecular weight of 24.0 kDa. The inhibitor was purified to near homogeneity with 181.55 fold purification and final yield of 29.18 per cent. The inhibitory potential of purified rice bean trypsin protease inhibitor on larvae of Spodoptera gut proteases was studied which showed maximum azocaseinolytic (1.93U), trypsin (0.66U) and chymotrypsin activity (0.37 U) in second larval instar which gradually decreased upto late fifth larval instar stage. Inhibitory assay revealed that rice bean trypsin inhibitors were strong inhibitor of Spodoptera litura and inhibited more than 80 per cent for trypsin and around 69 per cent for chymotrypsin activity. To further determine the efficacy of rice bean protease inhibitors, feeding assays were conducted by adding rice bean flour in larval artificial diet. High mortality rate was observed in Spodoptera litura larvae after 72 hrs when rice bean was given as sole diet (100% rice bean composition). The results obtained from present study provide important clues in designing strategies for sustainable use of rice bean protease inhibitors in developing insect-tolerant transgenic plants.

Prey preferences of notonectids towards larval mosquitoes across prey ontogeny and search area.

Predatory biological control agents can be effective natural means of managing pests, vectors and invasive species. However, the strength of predator-prey interactions can be regulated through context-dependencies that often remain unquantified. In particular, refuge effects can influence the efficacy of biological agents towards target species, and such effects are often driven by prey size and search area differences. In this study, we quantify the prey preferences of two predaceous notonectids, Anisops breddini and Anisops sardeus, towards four different aquatic larval instar stages of the medically important mosquito Culex quinquefasciatus across variations in surface area and water depth. Consumption rates differed significantly among the four larval sizes but not between the notonectids. Search area variations also elicited differences in consumption rates. Both predators tended to prefer second-instar mosquito prey among surface area and water depth variations, while generally avoiding the largest (fourth instar) and smallest (first instar) prey instar stages. For both predators, differential selectivity traits were emergent across surface area variations and water depth, with refuge effects for small prey generally greatest at intermediate-large depths with high surface areas. We thus demonstrate that predatory impacts of notonectids towards mosquito larvae differ significantly according to prey size, and likely peak at intermediate size classes. Different mosquito size classes often coexist and compete, selectivity has important implications for adult mosquito proliferations. Further, in ephemeral aquatic habitats where surface areas and water depths are highly variable spatiotemporally, the efficacy of notonectids in controlling mosquito prey may differ substantially. © 2019 Society of Chemical Industry.

Response of the bacterial community of Propylea japonica (Thunberg) to Cry2Ab protein

Propylea japonica is a very important predator in agricultural ecosystems, which could be exposed to Bt protein. In this study, the bacterial community of P. japonica fed with normal food and food containing Cry2Ab protein was characterized for the first time using qPCR and high-throughput sequencing approaches. Results showed no effect of Cry2Ab on P. japonica development and reproduction. The most abundant bacterial phylum was Firmicutes, and the most abundant genus was Staphylococcus. The total bacteria copy number was not significantly different across four larval stages. Bacteria species composition was gathered more closely in feed on sucrose solution (sucrose-fed) than in larvae only fed on pea aphid (aphid-fed), the diversity indices of some operational taxonomic unit (OTU) were significantly different between sucrose-fed and aphid-fed samples. Different instar larval stages of P. japonica fed with sucrose solution containing Cry2Ab Bt protein and found no effect on microbial community composition and total bacteria copy numbers. However, effects on relative abundance of microbes, copy numbers of Corynebacterium 1 and Glutamicibacter arilaitensis were observed significantly lower in Bt-fed first and fourth larval stages. Low and high concentrations of Cry2Ab protein altered the microbial abundance relative to sucrose-fed P. japonica and copy numbers of G. arilaitensis and Staphylococcus xylosus were significantly lower in Bt-fed samples than control sucrose-fed. Our results are the first report showing that feeding on Cry2Ab protein does not alter microbial species composition in P. japonica, but effects gene copy number of some dominant bacteria. Further investigations are needed to assess the effect of copy number change on P. japonica.

Gene expression during genital morphogenesis in the ground beetle Carabus maiyasanus.

To investigate the developmental genetics of genital formation in the carabid beetle Carabus maiyasanus, we compared gene expression patterns among five stages using transcriptomic RNA sequencing data from abdominal segments and genitalia in the third (last) larval instar (including prepupa) and pupal stages. We identified 18 839 genes, of which 10 796 were differentially expressed among stages or between sexes. There were relatively few differentially expressed genes (DEGs) between the sexes (3%). The DEGs were clustered into six groups, mainly according to stage-specific expression patterns. Genes in clusters 1-3 showed high expression levels before pupation and low expression levels during the pupal period, whereas genes in clusters 4-6 showed high expression levels from the prepupal to the pupal stages. Genes related to the initial pupation process and differentiation of genital discs in Drosophila were involved in clusters 4 and 6 and showed low expression levels at early third instar and elevated expression levels from the early prepupal stage, suggesting that the pupation process and genital differentiation started in the prepupal stage. Clusters 4 and 5 included developmental genes related to organ size control, which may be important in the formation of internal genital structures during the pupal stage.

Identification and Functional Analysis of a Delta Class Glutathione S-Transferase Gene Associated with Insecticide Detoxification in Bradysia odoriphaga.

A delta class glutathione S-transferase gene (BoGSTd2) is identified from Bradysia odoriphaga for the first time. Developmental expression analysis showed that expression of BoGSTd2 is significantly higher in the fourth instar larval stage and the adult stage. Tissue-specific expression analysis found that BoGSTd2 was expressed predominantly in the midgut and Malpighian tubules in the fourth instar larvae and the abdomen of adults. Expression of BoGSTd2 was significantly upregulated following exposure to chlorpyrifos and clothianidin. In vitro inhibition and metabolic assays indicated that recombinant BoGSTd2 could not directly metabolize chlorpyrifos and clothianidin. Nevertheless, disk diffusion assays indicated that BoGSTd2 plays an important role in protection against oxidative stress. RNAi assays showed that BoGSTd2 participates in the elimination of reactive oxygen species induced by chlorpyrifos and clothianidin. These results strongly suggest that BoGSTd2 plays an important role in chlorpyrifos and clothianidin detoxification in B. odoriphaga by protecting tissues from oxidative stress induced by these insecticides.

The steroid hormone 20-hydroxyecdysone binds to dopamine receptor to repress lepidopteran insect feeding and promote pupation.

Holometabolous insects stop feeding at the final larval instar stage and then undergo metamorphosis; however, the mechanism is unclear. In the present study, using the serious lepidopteran agricultural pest Helicoverpa armigera as a model, we revealed that 20-hydroxyecdysone (20E) binds to the dopamine receptor (DopEcR), a G protein-coupled receptor, to stop larval feeding and promote pupation. DopEcR was expressed in various tissues and its level increased during metamorphic molting under 20E regulation. The 20E titer was low during larval feeding stages and high during wandering stages. By contrast, the dopamine (DA) titer was high during larval feeding stages and low during the wandering stages. Injection of 20E or blocking dopamine receptors using the inhibitor flupentixol decreased larval food consumption and body weight. Knockdown of DopEcR repressed larval feeding, growth, and pupation. 20E, via DopEcR, promoted apoptosis; and DA, via DopEcR, induced cell proliferation. 20E opposed DA function by repressing DA-induced cell proliferation and AKT phosphorylation. 20E, via DopEcR, induced gene expression and a rapid increase in intracellular calcium ions and cAMP. 20E induced the interaction of DopEcR with G proteins αs and αq. 20E, via DopEcR, induced protein phosphorylation and binding of the EcRB1-USP1 transcription complex to the ecdysone response element. DopEcR could bind 20E inside the cell membrane or after being isolated from the cell membrane. Mutation of DopEcR decreased 20E binding levels and related cellular responses. 20E competed with DA to bind to DopEcR. The results of the present study suggested that 20E, via binding to DopEcR, arrests larval feeding and promotes pupation.

De Novo Variants in TAOK1 Cause Neurodevelopmental Disorders

De novo variants represent a significant cause of neurodevelopmental delay and intellectual disability. A genetic basis can be identified in only half of individuals who have neurodevelopmental disorders (NDDs); this indicates that additional causes need to be elucidated. We compared the frequency of de novo variants in patient-parent trios with (n = 2,030) versus without (n = 2,755) NDDs. We identified de novo variants in TAOK1 (thousand and one [TAO] amino acid kinase 1), which encodes the serine/threonine-protein kinase TAO1, in three individuals with NDDs but not in persons who did not have NDDs. Through further screening and the use of GeneMatcher, five additional individuals with NDDs were found to have de novo variants. All eight variants were absent from gnomAD (Genome Aggregation Database). The variant carriers shared a non-specific phenotype of developmental delay, and six individuals had additional muscular hypotonia. We established a fibroblast line of one mutation carrier, and we demonstrated that reduced mRNA levels of TAOK1 could be increased upon cycloheximide treatment. These results indicate nonsense-mediated mRNA decay. Further, there was neither detectable phosphorylated TAO1 kinase nor phosphorylated tau in these cells, and mitochondrial morphology was altered. Knockdown of the ortholog gene Tao1 (Tao, CG14217) in Drosophila resulted in delayed early development. The majority of the Tao1-knockdown flies did not survive beyond the third instar larval stage. When compared to control flies, Tao1 knockdown flies revealed changed morphology of the ventral nerve cord and the neuromuscular junctions as well as a decreased number of endings (boutons). Furthermore, mitochondria in mutant flies showed altered distribution and decreased size in axons of motor neurons. Thus, we provide compelling evidence that de novo variants in TAOK1 cause NDDs.

Laboratory evaluation of predation of Toxorhynchites amboinensis (Diptera:Culicidae) on three mosquito vectors of arboviruses in the Philippines

The spread of arboviral disease in the Province of Cavite, Philippines has resulted to the declaration of state of calamities which has led to higher disease burden and economic costs for the province and its population. With this situation, the use of biological control agents like the naturally occurring non - hematophagous Toxorhynchites species with predatory capacity at its larval stage was explored as an alternative approach to environmentally degrading chemical insecticides. Laboratory assessment of the predatory efficacy of wild type Toxorhynchites amboinensis was done independently against Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus prey larvae. Field collected Tx. amboinensis from selected areas of Cavite were reared until adult stage and allowed to oviposit in their rearing cages as a source of test insect. Using similar larval instar for the predator and prey species, predatory larvae were individually exposed to 20 and 40 prey densities in plastic cups. Predatory efficacy test was significantly different between the two prey densities offered. Higher predation rate ranging from 19.5 to 20.4 (48.75–51.00%) was recorded in the predatory larvae exposed to higher prey densities. Analysis of the predatory impact (PI) also revealed significant differences among the larval instars. The 4th instar larva was found to be the most predaceous which contributes a PI value of 19.74 to 20.08 (49.35–50.20%) at higher prey density with an overall PI of 15.75 to 17.68 (39.34–44.20%). The observed compulsive behavior of Tx. amboinensis at the latter part of the 4th larval instar stage can be considered as an additional positive behavior in reducing the larval stages of pest or vector population. This makes Tx. amboinensis a potential biological control agent and a potential component for inclusion in vector management program in the Province of Cavite.

Axon length maintenance and synapse integrity are regulated by c-AMP-dependent protein kinase A (PKA) during larval growth of the drosophila sensory neurons

Axonal extension and synaptic targeting are usually completed during early development, but the axonal length and synaptic integrity need to be actively maintained during later developmental stages and the adult life. Failure in the axonal length maintenance and the subsequent axonal degeneration have been associated with neurological disorders, but currently little is known about the genetic factors controlling this process. Here, we show that regulated intracellular levels of cAMP-dependent protein kinase A (PKA) are critical for the axon maintenance during the transition from the early to the later larval stages in the Drosophila class IV dendritic arborization (da) sensory neurons. Our data indicate that when the intracellular levels of PKA are increased via genetic manipulations, these peripheral neurons initially form synapses with wild-type appearance, at their predicted ventral nerve cord (VNC) target sites (in the first and second instar larval stages), but that their synapses disintegrate, and the axons retract and become fragmented in the subsequent larval stages (third larval stage). The affected axonal endings at the disintegrated synaptic sites still express the characteristic presynaptic and cytoskeletal markers such as Bruchpilot and Fascin, indicating that the synapse had been initially properly formed, but that it later lost its integrity. Finally, the phenotype is significantly more prominent in the axons of the neurons whose cell bodies are located in the posterior body segments. We propose that the reason for this is the fact that during the larval development the posterior neurons face a much greater challenge while trying to keep up with the fast-paced growth of the larval body, and that PKA is critical for this process. Our data reveal PKA as a novel factor in the axonal length and synapse integrity maintenance in sensory neurons. These results could be of help in understanding neurological disorders characterized by destabilized synapses.

Parental hypoxic exposure influences performance of offspring in Callosobruchus maculatus.

Modified atmosphere based on lack of O2 can protect stored grains from insect pest damage. Although population expansion of cowpea bruchid (Callosobruchus maculatus (Fabricius)) could be temporarily arrested when exposed to 2% O2 , this insect could survive extended periods of hypoxia and continue its normal development if normoxic conditions resumed. It is not clear whether parental hypoxic treatment has any effects on offspring performance and response to hypoxia. Hypoxia postponed development of treated parental bruchids at all stages. Its negative effects on oviposition and hatch rate of these eggs were significant only when hypoxia was administered at the parental fourth instar larval stage or later. When the F1 generation was exposed to hypoxia at the fourth instar larval stage, they exhibited comparable developmental delay and reduction in adult emergence and fecundity whether the parents experienced hypoxia or not. Interestingly, eggs laid by hypoxia-treated F1s had increased hatch rates if their parents had also been exposed to hypoxia. Stronger suppression of the digestive protease gene CatL and elevated basal expression of the stress responsive gene Hsp27 were observed in F1 larvae with parental hypoxic experience. Parental hypoxic experience appeared to better prepare the F1 progenies for further hypoxic challenge. © 2019 Society of Chemical Industry.

Impairment of pupation by RNA interference-aided knockdown of Broad-Complex gene in Leptinotarsa decemlineata (Say).

Dietary delivery of bacterially expressed double-stranded RNA (dsRNA) has a great potential for management of Leptinotarsa decemlineata. An important first step is to discover possible RNA-interference (RNAi)-target genes effective against larvae, especially the old larvae. In the present paper, five putative Broad-Complex (BrC) cDNAs (Z1-Z4, and Z6) were identified in L. decemlineata. The expression of the five LdBrC isoforms was suppressed by juvenile hormone signaling, whereas the transcription was upregulated by 20-hydroxyecdysone signaling at the fourth (final) instar larval stage. Feeding of bacterially expressed dsBrC (derived from a common fragment of the five LdBrC variants) in the third- and fourth-instar larvae successfully knocked down the target mRNAs. For the fourth-instar LdBrC RNAi hypomorphs, they had a higher larval mortality compared with the controls. Moreover, most dsBrC-fed beetles did not pupate normally. After removal of the apolysed larval cuticle, a miniature adult was found. The adult head, compound eyes, prothorax, mesothorax, metathorax were found on the dorsal view. Distinct adult cuticle pigmentation was seen on the prothorax. The mouthparts, forelegs, midlegs, and hindlegs could be observed on the ventral view of the miniature adults. For the third-instar LdBrC RNAi specimens, around 20% moribund beetles remained as prepupae and finally died. Therefore, LdBrC is among the most attractive candidate genes for RNAi to control the fourth-instar larvae in L. decemlineata.

Functional Response of Hyperaspis polita (Coleoptera, Coccinellidae) to the Recently Invaded Mealybug Phenacoccus solenopsis (Hemiptera, Pseudococcidae).

Hyperaspis polita Weise is the major predator of Phenacoccus solenopsis Tinsley in southwestern Iran. The functional response of fourth instar larva, adult female and male of the ladybird, H. polita, preying upon different stages of the mealybug P. solenopsis was evaluated. All stages of H. polita exhibited type II functional responses against all stages of P. solenopsis with an exception that the functional response of the adult female stage of H. polita to first instar P. solenopsis nymphs was type III. Changing prey and predator stages did not result in significant differences in attack rates. Handling times, however, differed for different prey and predator stages. This parameter was the shortest for any stage of predator fed on first instar P. solenopsis nymphs and was estimated to be 0.1100, 0.1868, and 0.2939h for fourth instar larval stage, adult females and males, respectively. Also, the handling time was the shortest for the fourth instar larval stage followed by adult females and males to different prey stages. Maximum predation rate (T/Th) decreased as P. solenopsis developed from one life stage to another, the lowest estimated value being obtained by feeding on adult female mealybug and the fourth instar larval stage was the most predatory. According to these results, the feeding potential of H. polita is noticeable with the fourth larval instar stage being considered the most efficient predatory stage for use in biological control programs. Further field-based studies are needed, however, before a comprehensive estimation of the biocontrol abilities of H. polita toward P. solenopsis can be made.

The Corazonin-PTTH Neuronal Axis Controls Systemic Body Growth by Regulating Basal Ecdysteroid Biosynthesis in  <i>Drosophila melanogaster</i>

Growth and maturation are coordinated by steroid hormone biosynthesis in various animal phyla. The insect steroid hormone, ecdysteroid, coordinates growth and maturation, represented by molting and metamorphosis. In Drosophila melanogaster, prothoracicotropic hormone (PTTH) neurons stimulate in generating peak levels of ecdysteroid to trigger maturation. In addition, recent studies have shed light on the role of PTTH signaling in basal ecdysteroid biosynthesis, which negatively affects systemic growth prior to maturation. However, it remains unclear how PTTH signaling is regulated for basal ecdysteroid biosynthesis. Here, we report that Corazonin (Crz)-producing neurons regulate basal ecdysteroid biosynthesis by affecting PTTH neurons. Inhibition of Crz neuronal activity increased pupal size, whereas it had little effect on pupariation timing. This phenotype resulted from enhanced growth and a delay in basal ecdysteroid elevation during the mid-third instar larval (L3) stage. Silencing of Crz in Crz neurons resulted in increased levels of the microRNA bantam, which represses basal ecdysteroid biosynthesis. Interestingly, Crz receptor (CrzR) expression in PTTH neurons was higher during the mid- than during the late-L3 stage. Silencing of CrzR in PTTH neurons increased pupal size, phenocopying the inhibition of Crz neuronal activity. When Crz neurons were optogenetically activated, a strong calcium response was observed in PTTH neurons in the mid-, but not the late-L3 stage. These data suggest that the Crz-PTTH neuronal axis modulates basal, but not peak ecdysteroid biosynthesis. Most significantly, this study uncovered a regulatory neuronal system affecting ecdysteroid biosynthesis in a developmental stage-specific manner.

Lifetables of Aproaerema modicella Deventer on soybean and soybean intercropped with pigeonpea

Field lifetables were prepared for determining mortality factors of leaf miner Aproaerema modicella Deventer on sole soybean and soybean intercropped with pigeonpea during kharif 2015. It was observed that the A. modicella completed three regular overlapping generations on sole soybean and as well on soybean intercropped with pigeonpea. Lifetable of field collected life stages revealed that late instar larval and pupal stages were the most vulnerable ones for mortality due to unknown reasons (predation and adverse climatic factors). The natural enemies viz., Apanteles sp. and Goniozus sp., also contributed some degree of mortality in larvae. Maximum larval parasitisation was observed with intercropped soybean. The negative values of trend index (<1) during the second and third generations showed that the mortality factors were effective in causing decline. The generation survival rate on sole soybean was 0.65, 0.70 and 0.63 which got reduced to 0.53, 0.69 and 0.62 on intercropped soybean, in the 1st, 2nd and 3rd generation, respectively. Thus, soybean intercropping with pigeonpea exhibited positive significant impact on key mortality factors of A. modicella.

Chemical composition of clove bud oil and development of clove bud oil loaded niosomes against three larvae species

Vesicular drug delivery systems have gained importance in new-age drug targeted therapies due to their simple and efficient mechanism. In this study, attempts were made on the synthesis of niosomes using clove bud oil and its evaluation against mosquito larvicidal activity. Using transmembrane pH method, clove bud oil loaded niosomes were prepared with cholesterol and surfactant variants of Span 20, 60 and 80. Dicetyl phosphate was added to avoid niosomes aggregation. The synthesized niosomes were characterized by Zeta potential, particle size, FT-IR and SEM analysis to evaluate charge, size, morphology and functional properties. Larvicidal property of niosomes was tested against II to IV instar larval stages of Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus at various concentrations for 24 h and LC50 value was calculated. The results exhibited that the clove bud oil loaded niosomes had greater larvicidal efficiency against tested mosquito species and these niosomes can be used as potential mosquito control agents. The results also suggested that niosome preparation could desirably be altered according to the drug of interest with greater stability and could be used in various biotechnological applications and drug delivery systems.

A Study of Cactoblastis cactorum (Lepidoptera: Pyralidae) in its Native Range: Further Insights into Life Cycle, Larval Identification, Developmental Parameters, Natural Enemies, and Damage to the Host Plant Opuntia ficus-indica (Caryophyllales: Cactaceae)

Cactoblastis cactorum Berg (Lepidoptera: Pyralidae) has been extensively studied since its initial use as a biological control agent for invasive populations of Opuntia Mill. The moth is native to several South American countries including Argentina where the exotic Opuntia ficus-indica (L.) Mill. (Cactaceae) is grown as a commercial crop. Recently C. cactorum has attracted considerable attention following its non-intentional establishment in Florida, because it now threatens the highly diverse and economically important Opuntia taxa of the southern USA and Mexico. To elucidate several aspects of this system, we recorded phenological data and parasitoid activity from Argentina across an annual cycle. We reared several generations of moths to better document the life cycle, described several formerly unpublished larval stages and morphological characters, and compared developmental parameters from samples collected from different sites. We found that C. cactorum has 3 overlapping generations across a 9 mo growth period with winter quiescence at the larval VI instar or pupal stage. The most common natural enemy of larvae was the parasitoid Apanteles opuntiarum (Martinez and Berta) (Hymenoptera: Braconidae). Information is given on its development and percentages of parasitism throughout the year. No egg parasitoids were found in field-collected eggsticks or on experimental eggsticks. There were no significant differences between developmental stages and times of C. cactorum from Tucuman and Cordoba in Argentina. We found intermediate C. cactorum damage on low-density cultivated Opuntia, but much lower damage in commercial plantations with high densities of plants. Surprisingly, we found that a “black spot” fungal infection (Alternaria Nees) (Pleosporaceae) produced a higher level of damage in commercial plantations in Cordoba, as well as in natural settings in Tucuman