Bee year: Basic physiological strategies to cope with seasonality.

Worker honey bees are subject to biochemical and physiological changes throughout the year. This study aimed to provide the reasons behind these fluctuations. The markers analysed included lipid, carbohydrate, and protein levels in the haemolymph; the activity of digestive enzymes in the midgut; the levels of adipokinetic hormone (AKH) in the bee central nervous system; the levels of vitellogenins in the bee venom and haemolymph; and the levels of melittin in the venom. The levels of all the main nutrients in the haemolymph peaked mostly within the period of maximal bee activity, whereas the activity of digestive enzymes mostly showed a two-peak course. Furthermore, the levels of AKHs fluctuated throughout the year, with modest but significant variations. These data suggest that the role of AKHs in bee energy metabolism is somewhat limited, and that bees rely more on available food and less on body deposits. Interestingly, the non-metabolic characteristics also fluctuated over the year. The vitellogenin peak reached its maximum in the haemolymph in winter, which is probably associated with the immunoprotection of long-lived winter bees. The analysis of bee venom showed the maximal levels of vitellogenin in autumn; however, it is not entirely clear why this is the case. Finally, melittin levels showed strong fluctuations, suggesting that seasonal control was unlikely.

Genome-wide identification of lipases in silkworm (Bombyx mori) and their spatio-temporal expression in larval midgut.

Lipases play crucial roles in food digestion by degrading dietary lipids into free fatty acids and glycerols. The domesticated silkworm (Bombyx mori) has been widely used as an important Lepidopteran model for decades. However, little is known about the lipase gene family in the silkworm, especially their hydrolytic activities as digestive enzymes. In this study, a total of 38 lipase genes were identified in the silkworm genome. Phylogenetic analysis indicated that they were divided into three major groups. Twelve lipases were confirmed to be expressed in the midgut at both transcriptional and translational levels. They were grouped into the same gene cluster, suggesting that they could have similar physiological functions. Quantitative real-time PCR (qRT-PCR) analyses indicated that lipases were mainly expressed in anterior and middle midgut regions, and their expression levels varied greatly along the length of midgut. A majority of lipases were down-regulated in the midgut when larvae stopped feeding. However, a unique lipase gene (Bmlip10583) showed low expression level during feeding stage, but it was significantly up-regulated during the larvae-pupae transition. These results demonstrated that expression of silkworm lipases was spatially and temporally regulated in the midgut during larval development. Taken together, our results provide a fundamental research of the lipase gene family in the silkworm.

Larval rearing and ontogeny of digestive enzyme activities in yellowfin seabream (Acanthopagrus latus, Houttuyn 1782).

The present research was conducted to provide insight into digestive larval capacity in Acanthopagrus latus larvae from hatching up to 30 days after hatching (DAH). Newly hatched larvae were stocked into six 300-L cylindrical polyethylene tanks at a density of larvae 50 larvae/L and reared by means of the green water system using Nannochloropsis oculata (0.5 × 106/mL). After mouth opening, larvae were fed with rotifers (5-16 individual/mL) from 2 to 20 DAH; then, Artemia nauplii (0.5-3.0 individuals/mL) were offered to larvae from 18 to 30 DAH, meanwhile a commercial microdiet was offered to larvae from 25 to 30 DAH. Larval performance in terms of growth and survival, and the assessment of the activity of selected digestive enzymes ontogeny of digestive enzymes activities was evaluated in larvae sampled at 0 (hatching), 7, 15, 22 and 30 DAH. Larvae showed an exponential growth characterized by two different growth stanzas, a first one characterized by slow growth rates comprised between hatching to 15 DAH (4.7 ± 0.2 mm), followed by a period of faster growth rates between 16 and 30 DAH (7.5 ± 0.6 mm). The activities of the brush border (alkaline phosphatase, ALP) and cytosolic (leucine-alanine peptidase, LAP) enzymes, as well as those of the pancreatic ones like total alkaline proteases, bile salt-activated lipase and α-amylase were detected from the mouth opening stage. Total activities of pancreatic and gastric enzymes increased with larval growth showing an enhancement of digestive capacities with larval age and size. The intestinal maturation in A. latus as assessed by the ratio of AP to LAP did not occur as expected by end of the first month of life suggesting the complete establishment of digestive luminal processes may take place at older ages. This study related to the growth patterns and ontogenic changes in activity of pancreatic, gastric and intestinal enzymes in A. latus and their nutritional regulation may be considered as the first step for improving the larviculture, as well as assessing and refining the nutritional requirements during the larval and early juvenile stages of this sparid species.

Implications of long-term exposure of a Lymantria dispar L. population to pollution for the response of larval midgut proteases and acid phosphatases to chronic cadmium treatment.

Cadmium (Cd) presence in terrestrial ecosystems is a serious threat that requires continuous development of biomonitoring tools. Ideally, a suitable biomarker of exposure should respond to the toxicant consistently in different populations regardless of previous exposure to pollution. Here we considered the activities and isoform patterns of certain proteases and acid phosphatases (ACP) in the midgut of Lymantria dispar larvae as well as the integrated biomarker response (IBR) for application in Cd biomonitoring. We compared the responses of caterpillars originating from unpolluted and polluted localities after they had been chronically subjected to dietary Cd (50 and 100 µg Cd/g dry food). The population inhabiting the unpolluted forest was far more sensitive to Cd exposure as the activities of total proteases, trypsin (TRY) and leucine aminopeptidase (LAP) were mostly reduced while the activities of total and non-lysosomal ACP were increased. Non-lysosomal ACP activity was elevated in larvae from the contaminated site in response to the higher Cd concentration. Exposure to the metal resulted in numerous alterations in the pattern of enzyme isoforms, but the responses of the two populations were similar except that larvae from the polluted locality were more tolerant to the lower Cd concentration. Non-lysosomal ACP activity and the appearance of ACP isoforms 4 and 5 together with the IBR index are the most promising indicators of Cd presence, potentially applicable even in populations with a history of exposure to pollution. TRY and total ACP activities could be used to monitor populations at uncontaminated localities.

Physiological and pathophysiological roles of acidic mammalian chitinase (CHIA) in multiple organs.

Acidic mammalian chitinase (CHIA) belongs to the 18-glycosidase family and is expressed in epithelial cells and certain immune cells (such as neutrophils and macrophages) in various organs. Under physiological conditions, as a hydrolase, CHIA can degrade chitin-containing pathogens, participate in Type 2 helper T (Th2)-mediated inflammation, and enhance innate and adaptive immunity to pathogen invasion. Under pathological conditions, such as rhinitis, ocular conjunctivitis, asthma, chronic atrophic gastritis, type 2 diabetes, and pulmonary interstitial fibrosis, CHIA expression is significantly changed. In addition, studies have shown that CHIA has an anti-apoptotic effect, promotes epithelial cell proliferation and maintains organ integrity, and these effects are not related to chitinase degradation. CHIA can also be used as a biomolecular marker in diseases such as chronic atrophic gastritis, dry eye, and acute kidney damage caused by sepsis. Analysis of the authoritative TCGA database shows that CHIA expression in gastric adenocarcinoma, liver cancer, renal clear cell carcinoma and other tumors is significantly downregulated compared with that in normal tissues, but the specific mechanism is unclear. This review is based on all surveys conducted to date and summarizes the expression patterns and functional diversity of CHIA in various organs. Understanding the physiological and pathophysiological relevance of CHIA in multiple organs opens new possibilities for disease treatment.

Effects of fluoranthene on digestive enzymes activity and relative growth rate of larvae of lepidopteran species, Lymantria dispar L. and Euproctis chrysorrhoea L.

Fluoranthene is one of the most abundant polycyclic aromatic hydrocarbon pollutants in the environment and it may accumulate in plant leaves which are the main food source for phytophagous insect species. The aim of this study was to establish the effects of dietary fluoranthene on specific activities of digestive enzymes and expression of their isoforms in the midgut, and the relative growth rates of Lymantria dispar and Euproctis chrysorrhoea larvae. Exposure to fluoranthene led to significantly decreased trypsin activity in the midgut of larvae of both species. Leucine aminopeptidase activity decreased significantly in the midgut of L. dispar larvae exposed to the lower concentration of fluoranthene, but that enzyme activity showed the opposite trend in E. chrysorrhoea larvae. There was no pollutant induced changes in lipase activity in L. dispar, while elevated enzyme activity was recorded in the midgut of E. chrysorrhoea larvae exposed to the lower concentration of fluoranthene. Different patterns of expression of enzyme isoforms were noticed. Relative growth rates of both species significantly decreased in fluoranthene treated larvae. These responses indicate to the significance of relationships between physiological changes and fitness-related traits in L. dispar and E. chrysorrhoea larvae affected by pollutant, and contribute to understanding the mechanisms of their adjustment to stressful conditions.

Estrogen-Related Receptor Influences the Hemolymph Glucose Content by Regulating Midgut Trehalase Gene Expression in the Last Instar Larvae of Bombyx mori.

The expression of trehalase in the midgut of insects plays an important role in glucose supply to the hemolymph. Energy metabolism is usually regulated by the estrogen-related receptor (ERR). A decrease in ATP levels is caused by the ERR hindering glycolysis. However, the relationship between trehalose accumulation and ERR expression is still unclear. Here, we found that silkworm ERR (BmERR) is concentrated and BmERR expression is strongly correlated with trehalase in the midgut during the last instar silkworm larval stage. We cloned the promoter of the trehalase from Bombyx mori (BmTreh) and found that the ERR bound directly to the core response elements of the promoter. Cell level interference and the overexpression of ERR can reduce or enhance BmTreh transcription and promoter activity. Overexpressed transgenic BmERR can significantly increase the expression of BmTreh in the midgut of the last instar silkworm larvae, thereby hydrolyzing trehalose into glucose and releasing it into the hemolymph. Additionally, increased hemolymph glucose content reduces silkworm pupa weight but does not affect silk protein production from the silk gland. Our results suggest a novel function for BmERR through its involvement in BmTreh regulation and expand the understanding of ERR functions in insect trehalose metabolism.

The Uncovered Function of the Drosophila GBA1a-Encoded Protein.

Human GBA1 encodes lysosomal acid ß-glucocerebrosidase (GCase), which hydrolyzes cleavage of the beta-glucosidic linkage of glucosylceramide (GlcCer). Mutations in this gene lead to reduced GCase activity, accumulation of glucosylceramide and glucosylsphingosine, and development of Gaucher disease (GD). Drosophila melanogaster has two GBA1 orthologs. Thus far, GBA1b was documented as a bone fide GCase-encoding gene, while the role of GBA1a encoded protein remained unclear. In the present study, we characterized a mutant variant of the fly GBA1a, which underwent ERAD and mildly activated the UPR machinery. RNA-seq analyses of homozygous mutant flies revealed upregulation of inflammation-associated as well as of cell-cycle related genes and reduction in programmed cell death (PCD)-associated genes, which was confirmed by qRT-PCR. We also observed compromised cell death in the midgut of homozygous larvae and a reduction in pupation. Our results strongly indicated that GBA1a-encoded protein plays a role in midgut maturation during larvae development.

Erucin modulates digestive enzyme release via crustacean cardioactive peptide in the elm leaf beetle Xanthogaleruca luteola (Coleoptera: Chrysomelidae).

Plant secondary metabolites influence the feeding in insects through several modes of action. In this study, the physiological effects of erucin isothiocyanate were investigated on the elm leaf beetleXanthogaleruca luteola(Müller) (Coleoptera: Chrysomelidae) via impact on crustacean cardioactive peptide (CCAP) and midgut digestive enzymes. Third instar larvae of elm leaf beetle were fed on leaves impregnated with erucin for three days. The results showed that erucin decreasedα-amylase, lipase, and protease release. Western blot analysis and competitive ELISA showed that erucin decreased CCAP content of the midgut, brain, and hemolymph. Moreover, incubation of dissected midgut with CCAP and also its injection into the hemocoel increased digestive enzyme release. It could be concluded that erucin isothiocyanate decreases CCAP content that itself led to a decrease in digestive enzyme release. Also, it suggests that CCAP could be one of the factors, regulating feeding activities in the elm leaf beetle. This report shows that CCAP is both a midgut factor and a neuropeptide that regulates digestive enzyme release in the elm leaf beetle and could be used to study erucin effects in insects.

Lacticaseibacillus casei in Diet of Juvenile Convict Cichlid Fish (Amatitlania nigrofasciata): Evaluating Growth Performance, Digestive Enzyme Activities, Immune Responses, and Stress Resistance.

In this study, growth performance, body composition, digestive enzymes activity, mucosal and immunological parameters, cultivable bacterial populations, and stress resistance were investigated in juvenile convict cichlid fish (Amatitlania nigrofasciata) that received a dietary supplement containing 0 (control), 5 × 106 (LC1), 5 × 107 (LC2), and 5 × 108 CFU g-1 diet (LC3) Lacticaseibacillus casei PB-LC39. Two hundred and forty fish (2.44 ± 0.04 g) were assigned to twelve experimental aquariums and feed ad libitum three times a day for 8 weeks. After final sampling, final body weight, percentage of weight gain (WG %), specific growth rate (SGR), food conversion ratio (FCR), and protein content of whole-body composition were significantly higher (P < 0.05) in fish fed LC1 diet than other treatments. Total protease, amylase, and lipase activities were also significantly higher (P < 0.05) in fish fed LC1 diet than other groups. Total protein from serum and mucus, lysozyme activity, total immunoglobulin (Ig), and serum globulin were significantly increased (P < 0.05) in fish fed LC1 diet compared with other groups. Moreover, total counts of lactic acid bacteria (LAB) in fish gut were significantly higher (P < 0.05) by different levels of L. casei PB-LC39 than the control group. Recovery rates of fish fed the probiotic cells, after an air-dive test, was significantly increased (P < 0.05) compared with the control group. Therefore, the results showed that L. casei PB-LC39 resulted in improving growth, health status, and stress resistance of fish during the rearing of juvenile convict cichlid fish.

Effects of graded levels of microbial fermented or enzymatically treated dried brewer's grains on growth, digestive and nutrient transporter genes expression and cost effectiveness in broiler chickens.

BACKGROUND: Poultry feed consists mainly of conventional grains and protein supplements, however, using treated unconventional agro-industrial by-products as replacements of corn soybean-based diet can minimize production costs and improve productivity. Therefore, in this study, the effects of fermented or enzymatically treated dried brewer grains (DBG) on growth, expression of digestive enzymes and nutrient transporters genes and the profitability of the rations were evaluated. A total of 1600 one-day-old Ross 308 broiler chicks were randomly distributed in 2 × 4 factorial arrangement (eight treatments with ten replicates, 20 birds/replicate). Experimental diets included two controls; negative control (basal corn-soybean diet; NC) and positive control (basal corn-soybean diet with exogenous enzymes; PC), and six diets in which basal diet was replaced by three levels of fermented DBG (FDBG; 5, 10 or 15%), or enzyme-treated DBG (DBG 5, 10 or 15%+Enz), for 38 days. RESULTS: The results described that feeding FDBG (three levels) or DBG5%+Enz improved (P < 0.05) BW gain and feed efficiency of broilers. Also, feeding FDBG10% yielded the best improvement in weight gain (10%), compared to NC group. Increasing the inclusion levels of DBG either fermented or enzymatically treated up-regulated (p < 0.01) expression of digestive-genes in proventriculus (PGC and PGA5, range 1.4-1.8 fold), pancreas (AMY2A, PNLIP, CELA1, and CCK; range 1.2-2.3 fold) and duodenum (CAT1, CAT2, GLUT1, GLUT2, LAT1, Pep1; range 1.3-3 fold) when compared to NC group. Feeding treated DBG significantly increased (p < 0.05, range 4.5-13.6%) gizzard relative weight compared to NC and PC groups. An additional benefit was lower (p < 0.01) cholesterol content from 66.9 mg/100 mg (NC) to 62.8 mg/100 mg (FDBG5 or 10%) in thigh meat. Furthermore, the least cost feed/kg body gain was achieved in FDBG10% and DBG5%+Enz groups, with approx. 16% reduction compared to NC cost, leading to increasing the income gross margin by 47% and 40% in FDBG10% and DBG5%+Enz groups, respectively. CONCLUSIONS: Substitution of corn-soybean based diet with 10% FDBG or 5% DBG+Enz resulted in better growth and higher economic efficiency of broilers chickens.

Evaluation of hydrolytic enzyme activities from digestive fluids of Anthonomus grandis (Coleoptera: Curculionidae).

The cotton boll weevil, Anthonomus grandis, is a major pest of cotton crops in South America. In this work, partial biochemical characterizations of (hemi) cellulases and pectinases activities in the digestive system (head- and gut- extracts) of A. grandis were evaluated. Gut extract section from third instar larvae exhibited endoglucanase, xylanase, ß-glucosidase, and pectinase activities. The endoglucanase and xylanase activities were localized in the foregut, whereas ß-glucosidase activity was mainly detected in the hindgut. In addition, no difference in pectinase activity was observed across the gut sections. Thus, A. grandis digestive system is a potentially interesting reservoir for further lignocellulolytic enzymes research.

Expression of the SARS-CoV-2 cell receptor gene ACE2 in a wide variety of human tissues.

BACKGROUND: Since its discovery in December 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected more than 2 180 000 people worldwide and has caused more than 150 000 deaths as of April 16, 2020. SARS-CoV-2, which is the virus causing coronavirus disease 2019 (COVID-19), uses the angiotensin-converting enzyme 2 (ACE2) as a cell receptor to invade human cells. Thus, ACE2 is the key to understanding the mechanism of SARS-CoV-2 infection. This study is to investigate the ACE2 expression in various human tissues in order to provide insights into the mechanism of SARS-CoV-2 infection. METHODS: We compared ACE2 expression levels across 31 normal human tissues between males and females and between younger (ages ≤ 49 years) and older (ages > 49 years) persons using two-sided Student's t test. We also investigated the correlations between ACE2 expression and immune signatures in various tissues using Pearson's correlation test. RESULTS: ACE2 expression levels were the highest in the small intestine, testis, kidneys, heart, thyroid, and adipose tissue, and were the lowest in the blood, spleen, bone marrow, brain, blood vessels, and muscle. ACE2 showed medium expression levels in the lungs, colon, liver, bladder, and adrenal gland. ACE2 was not differentially expressed between males and females or between younger and older persons in any tissue. In the skin, digestive system, brain, and blood vessels, ACE2 expression levels were positively associated with immune signatures in both males and females. In the thyroid and lungs, ACE2 expression levels were positively and negatively associated with immune signatures in males and females, respectively, and in the lungs they had a positive and a negative correlation in the older and younger groups, respectively. CONCLUSIONS: Our data indicate that SARS-CoV-2 may infect other tissues aside from the lungs and infect persons with different sexes, ages, and races equally. The different host immune responses to SARS-CoV-2 infection may partially explain why males and females, young and old persons infected with this virus have markedly distinct disease severity. This study provides new insights into the role of ACE2 in the SARS-CoV-2 pandemic.

Seasonal variations of digestive enzymes in sea cucumbers (Holothuria tubulosa, G. 1788) under culture conditions.

Sea cucumber, Holothuria tubulosa (Gmelin, 1788), is an economically valuable species due to its rich nutrients content that being exported in Turkey. However, culture of this species is difficult due to a lack of knowledge. The main objective of this study is to investigate seasonal activities of main digestive enzymes (protease, lipase, and amylase) for nutritional requirements. In this sense, sea cucumbers were stocked in the glass aquarium and sediment was 10 cm. It was replaced monthly and enriched (1% of the living individual weight) by dried Sargassum sp. Total protease and amylase enzymes have shown higher activity compared to lipase. All three enzyme activities were maximum in the summer and minimum in the winter. In summer, the highest measured total protease, amylase, and lipase activity values were 6.45 ± 0.66, 6.77 ± 0.72, and 2.78 ± 0.32 U/mg per protein, respectively. In winter, the lowest total protease, amylase, and lipase activity values were measured as 2.03 ± 0.16, 1.14 ± 0.14, and 0.12 ± 0.01 U/mg per protein, respectively. As a conclusion of the study, seasonal expression of the main digestive enzymes was strictly dependent on water temperatures and food abundance and also it was ideal to feed this species with food containing high protein and carbohydrate under appropriate temperatures for commercial culture.

Effect of 20-Hydroxyecdysone, a Phytoecdysteroid, on Development, Digestive, and Detoxification Enzyme Activities of Tribolium castaneum (Coleoptera: Tenebrionidae).

Plants present a delimited reservoir of biologically active compounds. Many plants synthesize several compounds of secondary metabolism, such as alkaloids, terpenoids, phenolics, steroids, etc. Such compounds are generally thought to be involved in plant-insect interactions. Phytoecdysteroids are a class of chemicals that plants synthesize; these compounds are analogues of molting hormones produced by insects. In this work, the effect of the 20-hydroxyecdysone, which is a molecule that belongs to the family of phytoecdysteroids, was tested on an insect pest, Tribolium castaneum (Herbst). Firstly, the effect of this molecule on post-embryonic development parameters was tested after ingestion at 300, 600, 900, and 1,200 ppm. Secondly, the effect of the 20-hydroxyecdysone was also tested on the biological parameters (proteins, alpha-amylase, detoxification enzymes). The results of the post-embryonic parameters test showed an important induction of larval mortality and a significant reduction of pupation and adult emergence rates. On the other hand, the test on the biological parameters showed that the 20-hydroxyecdysone caused a significant decrease in the levels of soluble proteins in treated larvae. In addition, the alpha-amylase activity was significantly inhibited by the ingestion of the phytoecdysteroid. And there was also a disruption of detoxification enzymes. The whole of the disturbances recorded in this work prove that phytoecdysteroids are thought to have potential value on T. castaneum control.

Feeding and digestion of the marine isopod Idotea emarginata challenged by poor food quality and microplastics.

Ingestion of microplastics can impair nutrition of marine invertebrates. In a laboratory study, we tested whether microplastics affect ingestion rates and gastrointestinal enzyme activities in the marine isopod Idotea emarginata. Isopods were fed for eight days with one out of four different food formulations: natural food (the brown alga Fucus vesiculosus) or synthetic diet consisting of freeze-dried algal powder embedded in agarose, both, with or without microplastic particles (fluorescent polymethyl methacrylate, 10-100 µm) at a concentration of 40 items per mg of food. The isopods accepted both types of food but consumed significantly more (average 3.1-fold) of the agar based synthetic food. I. emarginata responded to the reduced content of digestible organic matter in the synthetic food by a compensatory adjustment of the ingestion rates. Addition of microplastics had no effect on ingestion rates in natural food whereas the feeding rates for synthetic food varied in response to microplastics. Similarly, activity patterns of digestive enzymes, particularly those of esterases, changed significantly in the treatment with synthetic food. Isopods fed with synthetic food alone showed elevated esterase activities in the gut while those isopods fed with synthetic food and microplastics showed elevated esterase activities in the midgut gland but not in the gut. Apparently, not the exposure to microplastic alone, but the combined effects of reduced nutrient availability and microplastic ingestion caused considerable biochemical reactions in the digestive organs of the isopods.

Identification and enzymatic characterization of clip domain serine protease in the digestive fluid of the sea hare, Aplysia kurodai.

Clip domain serine proteases (CDSPs) participate in the extracellular signaling cascades of various biological processes such as innate immune responses in invertebrates. CDSP genes have been isolated from numerous invertebrates. Nevertheless, the enzymatic properties of mollusk CDSPs are poorly understood. In the present study, we demonstrated that the amino acid sequences of the trypsin-like serine protease purified from the digestive fluid of the sea hare, Aplysia kurodai resemble those of the unidentified CDSP-type protein (TPS3) of Aplysia californica predicted by genome analysis. The purified enzyme produced single 34 and 26.5 kDa bands on SDS-PAGE under non-reducing and reducing conditions, respectively. The 34-kDa band generated two amino-terminal sequences that were similar to the deduced sequences of the clip and catalytic domains of TPS3. The single amino-terminal sequence of the 26.5 kDa band showed a single sequence homologous to the catalytic domain. Thus, the purified enzyme consists of clip and catalytic domains bridged by disulfide linkage(s). The subsite specificity and inhibitor sensitivity of the purified enzyme were clearly distinct from those of horseshoe crab and silkworm CDSPs. A good substrate for the sea hare enzyme was pyroglutamyl-Arg-Thr-Lys-Arg-4-methyl-7-coumarylamide. The enzyme activity was strongly inhibited by aprotinin but not leupeptin. The physiological function of the enzyme in the digestive fluid remains to be determined.

Extracellular endonucleases in the midgut of Myzus persicae may limit the efficacy of orally delivered RNAi.

Myzus persicae is a major pest of many crops including canola and Brassica vegetables, partly because it vectors plant viruses. Previously it has been reported that double-stranded RNA delivered to aphids by injection, artificial diet or transgenic plants has knocked down target genes and caused phenotypic effects. While these studies suggest that RNA interference (RNAi) might be used to suppress aphid populations, none have shown effects sufficient for field control. The current study analyses the efficacy of dsRNA directed against previously reported gene-targets on Green peach aphid (Myzus persicae) strains. No silencing effect was observed when dsRNA was delivered in artificial diet with or without transfection reagents. dsRNA produced in planta also failed to induce significant RNAi in M. persicae. Transcriptome analyses of the midgut suggested other potential targets including the Ferritin heavy chain transcripts, but they also could not be knocked down with dsRNA. Here we show that dsRNA is rapidly degraded by midgut secretions of Myzus persicae. Analysis of the transcriptome of the M. persicae midgut revealed that an ortholog of RNases from other insects was abundant.

A differential transcriptional profile by Culex quinquefasciatus larvae resistant to Lysinibacillus sphaericus IAB59 highlights genes and pathways associated with the resistance phenotype.

BACKGROUND: The study of the mechanisms by which larvae of the Culex quinquefasciatus mosquito survive exposure to the entomopathogen Lysinibacillus sphaericus has benefited substantially from the generation of laboratory-selected colonies resistant to this bacterium. One such colony, RIAB59, was selected after regular long-term exposure of larvae to the L. sphaericus IAB59 strain. This strain is characterized by its ability to produce the well known Binary (Bin) toxin, and the recently characterized Cry48Aa/Cry49Aa toxin, able to kill Bin-resistant larvae. Resistance to Bin is associated with the depletion of its receptor, Cqm1 α-glucosidase, from the larvae midgut. This study aimed to identify novel molecules and pathways associated with survival of the RIAB59 larvae and the resistance phenotype. METHODS: A transcriptomic approach and bioinformatic tools were used to compare the profiles derived from the midguts of larvae resistant and susceptible to L. sphaericus IAB59. RESULTS: The RNA-seq profiles identified 1355 differentially expressed genes (DEGs), with 673 down- and 682 upregulated transcripts. One of the most downregulated DEGs was cqm1, which validates the approach. Other strongly downregulated mRNAs encode the enzyme pantetheinase, apolipoprotein D, lipases, heat-shock proteins and a number of lesser known and hypothetical polypeptides. Among the upregulated DEGs, the top most encodes a peroxisomal enzyme involved in lipid metabolism, while others encode enzymes associated with juvenile hormone synthesis, ion channels, DNA binding proteins and defense polypeptides. Further analyses confirmed a strong downregulation of several enzymes involved in lipid catabolism while the assignment of DEGs into metabolic pathways highlighted the upregulation of those related to DNA synthesis and maintenance, confirmed by their clustering into related protein networks. Several other pathways were also identified with mixed profiles of down- and upregulated transcripts. Quantitative RT-PCR confirmed the changes in levels seen for selected mRNAs. CONCLUSIONS: Our transcriptome-wide dataset revealed that the RIAB59 colony, found to be substantially more resistant to Bin than to the Cry48Aa/Cry49Aa toxin, developed a differential expression profile as well as metabolic features co-selected during the long-term adaptation to IAB59 and that are most likely linked to Bin resistance.

Effect of fluoranthene on antioxidative defense in different tissues of Lymantria dispar and Euproctis chrysorrhoea larvae.

This study examined the effect of long-term exposure to environmentally relevant concentrations of dietary fluoranthene (6.7 and 67 ng / g dry food weight) on defense mechanisms of the polyphagous forest insects Lymantria dispar L. and Euproctis chrysorrhoea L. The activities and expression of isoforms of superoxide dismutase (SOD) and catalase (CAT), the activities of glutathione S-transferase (GST) and glutathione reductase (GR), and total glutathione content (GSH) were determined in the whole midgut and midgut tissue, while SOD and CAT activities were assessed in hemolymph of the larvae. The results showed significant changes of enzyme activities, with more pronounced responses in larval midgut tissues, and between-species differences in patterns of response. Significantly increased activity of SOD was recorded in the whole midgut and midgut tissue of L. dispar larvae, as well as in midgut tissue of E. chrysorrhoea larvae. Fluoranthene increased CAT activity in midgut tissue of L. dispar larvae, and in the whole midgut and midgut tissue of E. chrysorrhoea larvae. Different expression patterns were detected for enzyme isoforms in tissues of larvae exposed to dietary fluoranthene. Total GSH content and GST activity increased in E. chrysorrhoea larval midgut tissue. Significantly decreased SOD activity in hemolymph of L. dispar larvae, and opposite changes in CAT activity were recorded in the hemolymph of larvae of two insect species. The tissue-specific responses of enzymes to dietary fluoranthene, recorded in each species, enabled the larvae to overcome the pollutant induced oxidative stress, and suggest further assessment of their possible use as early-warning signals of environmental pollution.