Midgut and fat body: Multisystemic action of pyriproxyfen on non-target organism Ceraeochrysa claveri (Neuroptera: Chrysopidae).

Morphological tools can assist in the evaluation of effects of insecticides on non-target insects. Pyriproxyfen, a juvenile hormone analog, is known to interfere with growth and metamorphosis of insects. However, there are studies showing indirect effects on natural enemies, including green lacewings. Few prior studies describe morphological effects of pyriproxyfen on target insect organs, especially on natural enemies. Through morphological tools, this study aimed to characterize the midgut and fat body, both important organs of digestion and great metabolic activity respectively, of the predator Ceraeochrysa claveri after chronic exposure to pyriproxyfen. Larvae of C. claveri were fed Diatraea saccharalis egg clusters treated with pyriproxyfen in solution of 50 or 100 mg a.i. L-1 throughout the larval stage. The biological data revealed significant increases in development time, especially in the third instar, and in cumulative mortality from the prepupal into the pupal stage. Morphological analysis of adult midgut (≤24 h old) showed damage including formation of epithelial folds, intercellular spaces, emission of cytoplasmic protrusions. Both fat body regions presented decrease of lipid droplets, vacuolization of trophocytes and mitochondrial injury featuring a multisystemic action. In both organs, pyriproxyfen exposure induced significant oxidative stress by mitochondrial superoxide production. Cytoprotective responses were induced in midgut and fat body cells by augmenting the number of cytoplasmic granules containing calcium and expression of HSP 90. Both organs proved to be efficient in presenting histopathological alterations, showing the sensitivity and applicability of this morphological tool for evaluating other insecticides in non-target organisms.

Pest and natural enemy: how the fat bodies of both the southern armyworm Spodoptera eridania and the predator Ceraeochrysa claveri react to azadirachtin exposure.

The effects of biopesticides on insects can be demonstrated by morphological and ultrastructural tools in ecotoxicological analysis. Azadirachtin-based products are widely used as biopesticides, affecting numerous insect populations. Through morphological biomarkers, this study aimed to characterize the fat bodies of both the southern armyworm Spodoptera eridania and the predator Ceraeochrysa claveri after chronic exposure to azadirachtin. Larvae of S. eridania and C. claveri were fed with fresh purple lettuce leaves (Lactuca sativa) and egg clusters of Diatraea saccharalis treated with azadirachtin solution of 6 mg active ingredient (a.i.)/L and 18 mg a.i./L for 7 days, respectively. The biological data showed a significant reduction in survival and body mass in S. eridania and cytotoxic effects in the parietal and perivisceral fat bodies in both species. Ultrastructural cell damage was observed in the trophocytes of both species such as dilated cisternae of the rough endoplasmic reticulum and swollen mitochondria. Trophocytes of S. eridania and C. claveri of the parietal and perivisceral layers responded to those injuries by different cytoprotective and detoxification means such as an increase in the amount of cytoplasmic granules containing calcium, expression of heat shock protein (HSP)70/HSP90, and development of the smooth endoplasmic reticulum. Despite all the different means of cytoprotection and detoxification, they were not sufficient to recover from all the cellular damages. Azadirachtin exhibited an excellent performance for the control of S. eridania and a moderate selectivity for the predator C. claveri, which presents better biological and cytoprotective responses to chronic exposure to azadirachtin.

Can exposure to neem oil affect the spermatogenesis of predator Ceraeochrysa claveri?

Novel biological control methods and integrated pest management strategies are basic requirements for the development of sustainable agriculture. As a result, there is a growing demand for research on the use of plant extracts and natural enemies such as the green lacewing, Ceraeochrysa claveri, as natural pest control methods. Studies have shown that although natural compounds such as neem oil (Azadirachta indica) are effective as pest control strategies, they also cause sublethal effects on nontarget insects, such as C. claveri. The aim of this study was to examine the effects of neem oil on C. claveri testes. C. claveri larvae were fed Diatraea saccharalis eggs, which were pretreated with 0.5%, 1%, and 2% neem oil. Testes were collected from larvae, pupae, and adults and analyzed using light and electron (transmission and scanning) microscopy. Changes in cellular stress and possible cell death were also determined by TUNEL assay and the marker HSP-70. The results showed that neem oil affects the organization and distribution of cysts in the testes and the normal sequence of cyst development, causing a delay in spermatogenesis in the testes of treated insects. Tests for cellular stress and DNA fragmentation indicated there was no cellular alteration in the treated groups. Although neem oil does not induce cell death or changes in HSP-70 expression, this biopesticide negatively impacts the process of spermatogenesis and could decrease the perpetuation of this species in the agroecosystem, indicating that the use of neem oil in association with green lacewings as a biological control should be carefully evaluated.

Neem oil (Azadirachta indica A. Juss) affects the ultrastructure of the midgut muscle of Ceraeochrysa claveri (Navás, 1911) (Neuroptera: Chrysopidae).

Cytomorphological changes, by means of ultrastructural analyses, have been used to determine the effects of the biopesticide neem oil on the muscle fibers of the midgut of the predator Ceraeochrysa claveri. Insects, throughout the larval period, were fed eggs of Diatraea saccharalis treated with neem oil at a concentration of 0.5%, 1% or 2%. In the adult stage, the midgut was collected from female insects at two stages of adulthood (newly emerged and at the start of oviposition) and processed for ultrastructural analyses. In the newly emerged insects obtained from neem oil treatments, muscle fibers showed a reduction of myofilaments as well as swollen mitochondria and an accumulation of membranous structures. Muscular fibers responded to those cellular injuries with the initiation of detoxification mechanisms, in which acid phosphatase activity was observed in large vesicles located at the periphery of the muscle fiber. At the start of oviposition in the neem oil treated insects, muscle fibers exhibited signs of degeneration, containing vacant areas in which contractile myofilaments were reduced or completely absent, and an accumulation of myelin structures, a dilatation of cisternae of sarcoplasmic reticulum, and mitochondrial swelling and cristolysis were observed. Enzymatic activity for acid phosphatase was present in large vesicles, indicating that mechanisms of lytic activity during the cell injury were utilized but insufficient for recovery from all the cellular damage. The results indicate that the visceral muscle layer is also the target of action of neem oil, and the cytotoxic effects observed may compromise the function of that organ.

Cytotoxic effects of neem oil in the midgut of the predator Ceraeochrysa claveri.

Studies of morphological and ultrastructural alterations in target organs have been useful for evaluating the sublethal effects of biopesticides regarded as safe for non-target organisms in ecotoxicological analyses. One of the most widely used biopesticides is neem oil, and its safety and compatibility with natural enemies have been further clarified through bioassays performed to analyze the effects of indirect exposure by the intake of poisoned prey. Thus, this study examined the cellular response of midgut epithelial cells of the adult lacewing, Ceraeochrysa claveri, to neem oil exposure via intake of neem oil-contaminated prey during the larval stage. C. claveri larvae were fed Diatraea saccharalis eggs treated with neem oil at concentrations of 0.5%, 1% and 2% throughout the larval stage. The adult females obtained from these treatments were used at two ages (newly emerged and at the start of oviposition) in morphological and ultrastructural analyses. Neem oil was found to cause pronounced cytotoxic effects in the adult midgut, such as cell dilation, emission of cytoplasmic protrusions, cell lysis, loss of integrity of the cell cortex, dilation of cisternae of the rough endoplasmic reticulum, swollen mitochondria, vesiculated appearance of the Golgi complex and dilated invaginations of the basal labyrinth. Epithelial cells responded to those injuries with various cytoprotective and detoxification mechanisms, including increases in cell proliferation, the number of calcium-containing cytoplasmic granules, and HSP 70 expression, autophagic processes and the development of smooth endoplasmic reticulum, but these mechanisms were insufficient for recovery from all of the cellular damage to the midgut. This study demonstrates that neem oil exposure impairs the midgut by causing sublethal effects that may affect the physiological functions of this organ, indicating the importance of studies of different life stages of this species and similar species to evaluate the safe and compatible integrated use of biopesticides.

Action of neem oil (Azadirachta indica A. Juss) on cocoon spinning in Ceraeochrysa claveri (Neuroptera: Chrysopidae).

Neem oil is a biopesticide that disturbs the endocrine and neuroendocrine systems of pests and may interfere with molting, metamorphosis and cocoon spinning. The cocoon serves protective functions for the pupa during metamorphosis, and these functions are dependent on cocoon structure. To assess the changes in cocoon spinning caused by neem oil ingestion, Ceraeochrysa claveri larvae, a common polyphagous predator, were fed with neem oil throughout the larval period. When treated with neem oil, changes were observed on the outer and inner surfaces of the C. claveri cocoon, such as decreased wall thickness and impaired ability to attach to a substrate. These negative effects may reduce the effectiveness of the mechanical and protective functions of cocoons during pupation, which makes the specimen more vulnerable to natural enemies and environmental factors.