Methanolic Extract of Winter Cherry Causes Morpho-Histological and Immunological Ailments in Mulberry Pyralid Glyphodes pyloalis.

The effect of Withania somnifera a medicinal plant seed extract was tested against lesser mulberry pyralid, a potential pest of mulberry. The mulberry leaves were used for silk production in rural areas of northern Iran. The extract was administered orally by leaf dipping method in two lower (5%W/V) and higher (15%W/V) dosages to third instar larvae (<24 h) for biological assays and to fifth instar larvae (<24 h) for Physiological studies. The results showed formation of larvoids (Ls), larval-pupal intermediates (LPIs), pupoids (Ps) and pupal-adult intermediates (PAIs). The results showed increased larval duration by 1.7 and 2 folds in 5 and 15% treatment, respectively. Fecundity of resultant adults was decreased by 1.2 and 1.3 in 5 and 15% treatment, respectively. Except approximate digestibility (AD) and consumption index (CI) all other feeding indices showed reduction. The feeding deterrence was prominent at 15% (87%) and 5% showing 48% deterrence. Our enzymatic and non-enzymatic assessments upon treatment showed reduction in key components, except detoxifying enzymes. However, the activity of an important enzyme involved in cuticle hardening and immunity called phenoloxidase was reduced. We also investigated the histology of midgut for further analysis and found drastic changes in main cellular elements. Immunological changes following treatment was noticeable in reduced Total Hemocyte Count but surprisingly increased Differential Hemocyte Count. However, the hemocytes structure was extremely damaged. The reduced number of eggs in treated but survived adults indicated reduced ovaries, with vacuolization both in trophocytes and oocytes. The key chemical compounds showed reductions particularly at 15%. The present results are concomitant with few earlier studies on this medicinal plant and deserve further studies particularly in deriving key chemicals that alter metamorphosis similar to insect growth regulators.

Characterization of a Digestive α-Amylase in the Midgut of Pieris brassicae L. (Lepidoptera: Pieridae).

The current study deals with a digestive α-amylase in the larvae of Pieris brassicae L. through purification, enzymatic characterization, gene expression, and in vivo effect of a specific inhibitor, Acarbose. Although α-amylase activity was the highest in the whole gut homogenate of larvae but compartmentalization of amylolytic activity showed an equal activity in posterior midgut (PM) and anterior midgut (AM). A three step purification using ammonium sulfate, Sepharyl G-100 and DEAE-Cellulose Fast flow revealed an enzyme with a specific activity of 5.18 U/mg, recovery of 13.20, purification fold of 19.25 and molecular weight of 88 kDa. The purified α-amylase had the highest activity at optimal pH and temperature of 8 and 35°C. Also, the enzyme had V max values of 4.64 and 3.02 U/mg protein and K m values of 1.37 and 1.74% using starch and glycogen as substrates, respectively. Different concentrations of acarbose, ethylenediamine tetraacetic acid, and ethylene glycol-bis (ß-aminoethylether) N, N, N', N'-tetraacetic acid significantly decreased activity of the purified α-amylase. The 4th instar larvae of P. brassicae were fed on the treated leaves of Raphanus sativus L. with 0.22 mM of Acarbose to find in vivo effects on nutritional indices, α-amylase activity, and gene expression. The significant differences were only found in conversion efficiency of digested food, relative growth rate, and metabolic cost of control and fed larvae on Acarbose. Also, amylolytic activity significantly decreased in the treated larvae by both biochemical and native-PAGE experiments. Results of RT-PCR revealed a gene with 621 bp length responsible for α-amylase expression that had 75% identity with Papilio xuthus and P. polytes. Finally, qRT-PCR revealed higher expression of α-amylase in control larvae compared to acarbose-fed ones.

Effects of an extracted lectin from Citrullus colocynthis L. (Cucurbitaceae) on survival, digestion and energy reserves of Ectomyelois ceratoniae Zeller (Lepidoptera: Pyralidae).

Lectins are the heterogeneous proteins in plants that serve as storage proteins via defensive mechanisms against herbivores. In the current study, a lectin was extracted and purified from seeds of Citrullus colocynthis by Sepharose 4B-Galactose and DEAE-cellulose fast flow chromatographies. Different concentrations of the lectin were added to artificial diet of Ectomyelois ceratoniae larvae finding out its effect on some biological parameters, digestive physiology and amount of storage macromolecules. It was found that CCA (C. colocynthis Agglutinin) increased life span from 23.44 days in control to 28.59 days in the treated individuals. Survival of larvae on control and CCA diets were 93.3 and 66.6%, respectively. Different concentrations of CCA significantly affected α-amylase and general proteolytic activities except for TAG-lipase activity. Activities of all specific proteases decreased when larvae were fed on different concentrations of CCA except for aminopeptidase. Meanwhile, amount of storage macromolecules in the larvae fed on different concentrations of CCA statistically decreased vs. control. These results demonstrated that CCA could intervene in physiology of E. ceratoniae and survival of larvae. Therefore, it can be taken into consideration in IPM of the pest through plant breeding programs.

Effect of milk thistle, Silybium marianum, extract on toxicity, development, nutrition, and enzyme activities of the small white butterfly, Pieris rapae.

The methanolic extract of milk thistle, Silybium marianum L. (Asterales: Asteraceae), was investigated for its effects on the mortality, growth, feeding indices, enzymatic activity, and levels of non-enzymatic molecules of the small white butterfly, Pieris rapae L. (Lepidoptera: Pieridae), a pest of cruciferous plants. Feeding indices including approximate digestibility (AD), efficiency of conversion of digested food (ECD), efficiency of conversion of ingested food (ECI), relative growth rate (RGR), and relative consumption rate (RCR) were measured. These indices were variously affected: the RGR, RCR, and AD decreased, but the ECD and ECI increased. The LC50 and LC25 values were estimated as 2.94% and 1.20%, respectively. At the lowest concentration of S. marianum extract (0.625%), the feeding deterrence index was 40.48%. The duration of the pupal stage and the rate of larval growth decreased. These changes may be due to alterations in metabolic activity, such as the increase in alkaline phosphatase activity, which is likely involved in detoxification. Additionally, the activities of alanine aminotransferase and aspartate aminotransferase, which are key components of amino acid catabolism, decreased. The amount of glucose (an energy source) and uric acid (the excreted end product) increased, while total protein (another energy source) and cholesterol decreased. These results indicate that this plant possesses potential secondary metabolites that may be useful for the future study of the control of insect pests.